EPIDEMIOLOGY AND MANAGEMENT OF COTTON LEAFROLL DWARF VIRUS, AN EMERGING INSECT-TRANSMITTED VIRUS IN THE SOUTHERN U.S

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: OTHER GRANTS
• Reporting Frequency: Annual
• Accession No.: 1021180
• Grant No.: 2019-70006-30441
• Cumulative Award Amt.: $323,661.00
• Proposal No.: 2019-02957
• Project Start Date: Sep 1, 2019
• Project End Date: Aug 31, 2022
• Grant Year: 2019
• Program Code: [ARDP]- Applied Research and Development Program

Recipient Organization
AUBURN UNIVERSITY
108 M. WHITE SMITH HALL
AUBURN,AL 36849

Performing Department
Entomology & Plant Pathology

Non Technical Summary
Cotton blue disease (CBD), caused by the aphid-transmitted Cotton leafroll dwarf virus (CLRDV) (genus Polerovirus, family Luteoviridae), is an emerging disease that poses a significant threat to the profitability of the U.S. cotton industry.CLRDV is regarded as the second most damaging virus disease to commercial cotton worldwide and is the first virus reported to cause yield loss in the southeastern U.S. CLRDV was initially identified in symptomatic cotton collected from two Alabama counties in 2017. In 2017, CBD incidence and yield loss were estimated for approximately 50,585ha of cotton in south AL, and yield loss was estimated at an average of 560 kg/ha across this area, which was valued at $19 million dollars. In 2018, late-season surveys detected this virus in 23 AL counties, 14 GA counties, 13 MS counties, FL, and SC. In 2018, visual surveys of symptoms in south AL ranged from negligible to near 100% incidence, although incidences of 10-20% were more commonly observed in GA and MS. A severely infected field in south AL had ~100% square drop. Although information on the geographic range, symptoms and incidence are preliminary theydemonstrate the potential for this disease to reduce plant health and yield. This multidisciplinary research project will begin addressing agronomics, crop genetics, and insect pest management components of cotton production impacting CBD severity. Knowledge will be generated about aphid population dynamics responsible for virus spread in the landscape, the efficacy of currently available management practices to reduce virus incidence and severity, and will support cotton breeding efforts to develop varieties that are resistant to CBD. Research conducted for this project will ultimately result in interdisciplinary management approaches for this emerging problem by providing valuable knowledge, training tools, and data that will benefit stakeholders in in all areas wherethisdiseaseis detected.

Animal Health Component

25%

Research Effort Categories

Basic

50%

Applied

25%

Developmental

25%

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
212	1710	1170	80%
216	1710	1130	20%

Knowledge Area
212 - Pathogens and Nematodes Affecting Plants; 216 - Integrated Pest Management Systems;

Subject Of Investigation
1710 - Upland cotton;

Field Of Science
1130 - Entomology and acarology; 1170 - Epidemiology;

Keywords

cotton leafroll dwarf virus

cotton blue disease

aphid dispersal

aphid transmitted virus

Goals / Objectives
This multidisciplinary research project will begin addressing agronomics, crop genetics, and insect pest management components of cotton production impacting CBD severity, and will ultimately result in interdisciplinary management approaches for this emerging problem. Research conducted for this project will provide valuable knowledge, training tools, and data that will benefit stakeholders in in all areas where CLRDV is detected. The specific objectives of this project are to: 1) Characterize aphid population dynamics in cotton agroecosystems and identify the timing of primary spread of CLRDV to cotton; 2) Evaluate the efficacy of host plant resistance, vector management and cultural practices on reducing CBD incidence, disease severity and yield loss; 3) Develop educational materials and programs to increase stakeholder knowledge and adoption of an integrated pest management program for managing CBD. We will monitor aphid dispersal and colonization events, sentinel plants, and CBD symptom severity and incidence weekly throughout the cotton production season in small plot replicated trials designed to quantify the impact of planting date, chemical management of vector populations, and host plant resistance on reducing yield losses caused by CBD. Specifically, the results from this study will generate foundational knowledge about aphid-vector population biology and epidemiology of CLRDV in the U.S., and will inform efforts to devise effective integrated management practices for CBD that minimize economic, environmental and human health risks. Project outcomes will be disseminated to scientific and stakeholder communities to increase awareness of CBD and adoption of IPM practices.This project meets the goals of the IPM roadmap by devising integrated disease management strategies that prevent unacceptable levels of CBD damage, reduce the evolution of resistance to pesticides in vector populations, and minimize economic, environmental, and human health impacts. The research conducted under this project will advance fundamental knowledge about epidemiological processes that underlie virus spread and incidence, and determine the efficacy of cultural and chemical management tools for reducing disease-associated losses. The synthesis of these objectives will inform CBD management strategies, facilitate adoption of management approaches through grower-oriented extension meetings and online materials, and will promote coordination of research and extension efforts across the southeastern cotton production region affected by CBD. All of the epidemiological processes and management practices investigated in this proposal are relevant for other cotton producing regions, and can be used to guide management and research efforts in other areas of the U.S. cotton belt where CBD is detected. Identification of resistance in cotton will also have broad impacts on U.S. cotton breeding programs across the public and private sectors.

Project Methods
Objective 1. Characterize aphid population dynamics in cotton agroecosystems and identify the timing of primary spread of CLRDV in cotton (Research Objective). In the fields used for small plot replicated trials (Obj. 2), beginning the week of planting and continuing until the end of August, dispersing aphids, colonizing aphids, and sentinel plants will be monitored at two locations in AL and two locations in GA. Yellow tile pan traps will be used to attract and capture immigrating aphids around cotton fields using previously described methods. Healthy, two-true leaf cotton that was maintained in an insect and virus-free greenhouse will be used as a sentinel plant that will be attractive to aphids and susceptible to virus infection (grown from untreated DP 1646 B2XF cottonseed). A minimum of four pan traps will be positioned around each field and aphids captured in traps will be collected weekly and stored in a vial of 70% ethanol until they are slide mounted and identified. A minimum of eight sentinel plants, each planted in a 6" pot, will be placed around the field margin, spaced equidistant along the edges. Sentinel plants will be replaced weekly. Sentinel plants will then be sprayed with an insecticide and maintained in an insect-free greenhouse (sprayed weekly with an insecticide) for a period of 30 days. Aphid abundance and species composition from the pan traps and sentinel plants will be recorded for each week of trapping. The sentinel plants will be monitored in the greenhouse for the presence/absence of symptoms on each plant. After 30 days they will be tested for infection with CLRDV. Timing of virus spread will be compared among locations, and the incidence of infection in sentinel plants will be compared among field locations. Data will also be analyzed to test for correlations between specific aphid species and proportion of sentinel plants infected across trapping periods and locations.Objective 2. Evaluate the efficacy of management tools and strategies for reducing CLRDV incidence, disease severity and yield loss (Research and Extension Objective). HPR, management of aphid vectors, and planting date will be evaluated in small plot replicated field experiments conducted at two locations in each state to quantify their impact on reducing disease incidence, severity and yield loss. Four insect management regimes will be evaluated on two different planting dates to determine if aphid management will reduce final incidence of CLRDV. Aphid populations resulting from natural infestations will be manipulated with insecticide applications to examine population dynamics in cotton related to colonization, spread within the crop, symptom severity, final virus incidence, yield, and fiber quality. The four insecticide regimes compared in this objective include: 1.) No management of cotton aphid; 2.) Attempt to prevent colonization of the crop making weekly applications of insecticides beginning at the 1-true-leaf stage; 3.) Spray after colonizing aphids are found to prevent population buildup and secondary spread in the crop. After first detection, make weekly applications of insecticides to deter subsequent colonization events. 4.) Make a calendar-based application of insecticides the first week of July (per current grower practices), and then weekly applications of insecticides to deter subsequent colonization events. Stand counts will be taken during the seedling stage in rows 2 & 3 of each plot by counting the total number of plants in each row. All plots will be scouted weekly for aphids throughout the growing season by visually examining plants in the middle two rows of each plot. Each week, aphids counts will be performed on ten randomly selected plants in the middle two rows of each plot. The total number of live alate and apterous individuals will be recorded separately on one upper, middle and lower fully expanded leaf of each plant. Leaves will be selected randomly from the terminal leaf of branches. In addition, ten consecutive plants in each of rows 2 and 3 will be visually inspected for aphids, and the presence/absence of aphids will be recorded to determine the proportion of plants infested with at least one aphid per plant during each evaluation period. CLRDV symptoms present in the middle two rows of each plot will also be monitored weekly on these the same ten consecutive plants. During each evaluation, the presence/absence of the different symptoms will be recorded. Plant-growth related symptomology that will be recorded includes leaf crinkling, cupping and thickening of leaves, reddening of leaf veins and petioles, shortened internodes, dwarfing of the plant, swollen and brittle stems, and accentuated verticality of top growth. Samples of plants exhibiting each symptom type will be collected and tested for virus to confirm that symptoms observed are associated with the virus.Additional measurements will be taken after defoliation to further document symptomology and yield effects. Comprehensive plant mapping will be conducted on 10 randomly sampled plants per plot to assess the fruiting profile. All first and second positions will be mapped starting at the lowest fruiting position, and the plant height, total number of nodes, and presence of shortened internodes will be recorded. The two middle rows of each plot will be harvested using a 2-row spindle picker and the resulting seedcotton will be collected in individual bags, weighed and submitted to the UGA Microgin for ginning and subsequent fiber quality analyses. These will include measurements of changes in fiber color, leaf grade, fiber length, micronaire, fiber strength, length uniformity index, and trash percent.Objective 2b. Host plant resistance: two experiments will be conducted. The first will phenotype germplasm containing resistance genes to previously reported poleroviruses for their susceptibility to U.S. strains of CLRDV in greenhouse experiments. The second will phenotype susceptibility to CBD in small plot replicated trials. A large-scale field screening trial will have a minimum of 500 germplasm lines from diverse origins for resistance to the strain of CLRDV in the southern U.S. The lines to be tested will include elite germplasm recently developed by the public breeding programs, obsolete cultivars, and non-adapted germplasm such as landrace accessions in the U.S. National Germplasm Collection. These trials will be monitored for aphids and virus.Objective 3: Develop educational materials and programs to increase stakeholder knowledge and facilitate the adoption of an integrated pest management program for managing CBD (Extension Objective):Once the data collection are completed each year, research faculty will analyze and discuss interpretation of the data with Extension specialists. Together, these faculty members will develop outreach slides and news releases that can be extended to growers through existing conduits. Cotton team Extension specialists in Georgia and Alabama maintain close ties with growers and conduct annual county level production meetings, and participate in annual Georgia and Alabama Consultants Association continuing education workshops, as well as field days. In addition, Extension fact sheets, popular press articles, postings on the UGA Cotton Team blog, and peer-reviewed articles will be written about research findings. Broader scientific and stakeholder communities will be targeted through Extension bulletin repositories, the Plant Management network, leading farm publications, and popular press outlets. The impact of outreach efforts in reaching key stakeholders will be conducted using the impact evaluation unit at UGA that will work with project leaders from the idea stage of project development, serving as a member of the management team to develop the evaluation plan based on the Request for Application requirements, project goals, and objectives.

Progress 09/01/19 to 08/31/22

Outputs
Target Audience:The target audiences of these projects include the research community, university extension entomologists, Cotton Inc, The National Cotton Council, county and regional extension agents, crop consultants, and producers in the Southern U.S. who are managing cotton. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Co-PIs, Students, and postdocs have presented research related to CLRDVat virtual national society meetings during the past fiscal year. PI and co-PI participation in regional working group meetings have enabled us to begin coordinating regional research efforts in entomology, plant pathology, and plant breeding to expand research efforts beyond the scope of this grant and get similar data on symptomatology, aphid dispersal, and varietal differences in susceptibility across the southeast and mid-south. How have the results been disseminated to communities of interest?The current state of knowledge about CLRDV distribution, incidence, spread, and management has been disseminated at local and regional extension meetings, annual working group meetings, and annual meetings of professional societies (including, the Entomological Society of America, and Annual Phytopathological Society, the Beltwide Cotton Conferences, and the National Conservation Systems Cotton and Rice Conference). Preliminary results along with diagnostic guides with images have been disseminated to stakeholders via articles in print and electronic media, and social media outlets. Information shared at these venues and in extension publications has been disseminated by several popular media outlets, including Farm Press and local media. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Objective 1: Characterize aphid population dynamics in cotton agroecosystems and identify the timing of primary spread of CLRDV to cotton Aphids were trapped in three distinct cotton production regions in north, central, and south Alabama. These locations vary in climate, soil type, and native vegetation and represent analogous production regions in other southeastern states on the eastern shore board. Matching funds from Cotton Inc. (included as a match for this project, and additional funds not originally included in the matching funds) provided support for entomologists from VA, NC, SC, GA, FL, MS, and TX to monitor for aphid dispersal at one location during the cotton growing season. Aphids were trapped/monitored every week for 2-years., their total numbers were counted, and eight species reported to feed on cotton were identified to species and enumerated. Concurrent with insect trapping at the Alabama locations, healthy two-true leaf cotton were used as sentinel plants weekly from March - November to detect weeks that CLRDV spread was occurring in the landscape. The insect identification was completed early summer of 2022. During the summer of 2022 a draft identification guide for cotton-infesting aphids monitored using yellow pan traps has been completed and will be submitted as an extension guide in the Fall of 2022. All other data has been entered and a graduate student is conducting analyses to examine seasonal population trends, aphid species composition, and abundance among locations. A manuscript draft will be submitted for publication in 2023 after the analyses are finalized. Preliminary results from the regional aphid trapping show differences in the abundance of cotton aphids, the reported vector of CLRDV. More cotton aphids are found in south AL, and GA where higher incidence of CLRDV has been reported, whereas, numbers were much lower in states where low incidence of CLRDV has been reported. Seasonal peaks of cotton aphid populations are also variable among locations. In AL, where CLRDV spread was monitored for 33 weeks, only one week of virus spread was detected at the north AL location, 11 weeks of virus spread were detected in central AL, and 16 weeks of virus spread were detected in south AL. Virus spread occurred as early as April (pre-plant), during the summer months, and as late as November (after harvest). It also occurred at times when cotton aphids were not the predominant species, suggesting other vectors may be present in the landscape. The results of these studies are characterizing seasonal dispersal patterns of the vector and virus in the landscape, and are helping to characterize the epidemiology of CLRDV. The regional differences may help to explain the variation in the distribution and incidence of CLRDV across the landscape, and may relate it to the abundance and distribution of vector populations. Objective 2:Evaluate the efficacy of host plant resistance, vector management, and cultural practices in reducing CBD incidence, disease severity, and yield loss Replicated field experiments were conducted in AL and GA to determine whether or not managing seasonal infestations of the vector of CLRDV, the cotton aphid, can reduce the final incidence of CLRDV. Pesticide spray regimes that ranged from weekly sprays to one spray that is more typical of current management practices were examined. CLRDV incidence was quantified using RT-PCR to detect the virus in randomly collected samples from each experimental plot. These results showed that aphid management was ineffective at reducing virus incidence. Final incidence ranged from 60-100% in all small plots examined. The results of this study were published in Crop Protection in 2022. This information shows the importance of identifying alternative management strategies for CLRDV. Another field experiment was conducted in Alabama over a three-year period to examine the interaction between the timing of CLRDV inoculation and plant growth stage on final yield. Small plots were caged to exclude natural populations of aphids so that insect infestations could be experimentally controlled. Plots were infested with aphids carrying the virus either 1, 2, 3, and 4-weeks after planting; non-infested plots were used as controls. In one year yield was generally associated with the timing of infection with the lowest yield observed in the plots that were infested 1 week after emergence, and the highest yield observed when the plots were not infested. The second and third year there was no treatment effect. Weather station data collected during these experiments show very different temperatures and soil moisture among years. Overall, these results suggest that younger crops may be more susceptible to CLRDV-related yield loss, but additional abiotic stress may play a role in yield loss outcomes. A draft manuscript of these results is being finalized and will be submitted for publication fall 2022. Host plant resistance was examined using matching funds provided by Cotton Inc. At Auburn University, plant breeding efforts have been focused on identifying resistance in the form of immunity to CLRDV in cotton germplasm. To date, over 2000 advanced breeding lines, obsolete varieties, and commercial cultivars have been assessed with 5 genotypes giving negative PCR tests for CLRDV in two years. A leaf disk assay confirmed that all germplasm can be infected with the virus. Therefore it was identified that none of the genotypes were immune to infection. It was noted that genotypes do respond differently to the virus in regard to symptom expression. This is important in evaluating tolerance to the virus. In addition, screening techniques for grafting were evaluated and published. This is important as there was not a detailed guide on how to screen for viruses in cotton through grafting. At UGA marker-trait association studies are being used to identify putative resistance genes in two populations that are segregated for viral symptoms. These ongoing efforts are seeking to identify resistance traits that can be bred into elite cotton lines because resistant varieties are the best way to manage plant viruses. Objective 3: Develop educational materials and programs to increase stakeholder knowledge and adoption of an integrated pest management program for managing CBD The information generated from these research studies has been communicated to agricultural stakeholders and scientists at extension venues and scientific conferences. Results are shared with Land Grant University Extension and Research Faculty at annual meetings of the Southern States Cotton Arthropod Working Group (14 states), and the Southeastern Row Crop Entomology Working Group (six states) members. In GA information about CLRDV was disseminated to a total of 709 attendees at 18 cotton production meetings, approx.. 100 extension agents at county agent trainings, and to producer and industry stakeholders at the Georgia Cotton Commission Research Field Day and the UGA Cotton and Peanut Research Field Day. In AL cotton disease updates were presented at 7 Alabama Crops Tours. In addition, invited updates on CLRDV were presented as a "Focus on Cotton: Cotton Leafroll Dwarf Virus in the U.S." through the American Phytopathological Society's Grow Plant Health Exchange Webinar, and at the Southern IPM Hour Webinar hosted by the Southern IPM Center in 2022. Two extension publications have also been produced, one in AL and one in GA: Hand, C. and UGA Extension Cotton Team. 2022. Georgia Cotton Production Guide. University of Georgia Annual Publication 124-2. Found online at https://secure.caes.uga.edu/extension/publications/files/pdf/AP%20124-2_1.PDF Conner, K. N.; Strayer-Scherer, A.; Hagan, A. K.; Koebernick, J.; Jacobson, A.; Bowen, K. L.; Sikora, E. J.; Graham, S.; and Brown, S. 2021. Cotton leafroll dwarf virus. ANR-2539. https://www.aces.edu/blog/topics/crop-production/cotton-leafroll-dwarf-virus/

Publications

• Type: Journal Articles Status: Published Year Published: 2022 Citation: Mahas, J.W., F.B. Hamilton, P.M. Roberts, C.H. Ray, G.L. Miller, M. Sharman, K. Conner, S. Bag, E.K. Blythe, M. D. Toews, A.L. Jacobson. Investigating the effects of planting date and Aphis gossypii management on reducing the final incidence of cotton leafroll dwarf virus. Crop Prot. 158: 106005.
• Type: Journal Articles Status: Published Year Published: 2022 Citation: Conner, K. N., Sikora, E., Koebernick, J., & Zaccaron, M. (2022). Interdisciplinary Team Addresses Cotton Leafroll Dwarf Virus in Alabama. The Journal of Extension, 60(2), Article 9. https://doi.org/10.34068/ joe.60.02.09
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Roberts, P. and S. Bag. 2022. Impact of aphid insecticide timing on cotton yield and cotton leafroll dwarf virus. 2022 Beltwide Cotton Conferences. San Antonio TX.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: *Mahas, J.W., A.L. Jacobson, C. Ray, J.B. Mahas, M. Toews, P. Roberts, J. Greene, G.G. Kennedy, S. Taylor, S. Paula-Moraes, J. Gore, W. Crow, S.D. Stewart, D. Kerns and S. Vyavhare. 2022. Monitoring Seasonal Dynamics and Community Composition of Cotton Infesting Aphid Species in the Southern U.S. Beltwide Cotton Conferences, San Antonio TX. Oral presentation.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Roark, D., J. Koebernick and A.L. Jacobson 2022. Protocol for Screening for Aphid Resistance in Cotton. Beltwide Cotton Conferences, San Antonio TX. Poster presentation.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: *Mahas, J.B., J.W. Mahas, C. Ray, A. Kesheimer, K. Conner and A.L. Jacobson. 2022. Monitoring the Seasonal Occurrence of Cotton Leafroll Dwarf Virus Spread in Relation to Aphid Vector Flights in Alabama. Beltwide Cotton Conferences, San Antonio TX. Oral presentation.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: *Schlarbaum, K., K. Conner, A. Kesheimer, A.L. Jacobson. 2022. Investigating the Interaction of Plant Age and Timing of Cotton Leafroll Dwarf Virus Infection on Yield Loss. Beltwide Cotton Conferences, San Antonio TX. Oral presentation.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: *Mahas, J.W., J. Ahl, C. Ray, Southeastern Row Crop Entomology Working Group, Midsouth Row Crop Entomology Working Group, A.L. Jacobson. 2021. Monitoring cotton infesting aphid species in the southern U.S. National Meeting of the Entomological Society of America. 10 minute oral presentation. November 2021.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Pandey, S., S. Bag, P. Roberts, A.L. Jacobson, R. Srinivasan. 2021. Aphid-mediated transmission of cotton leafroll dwarf virus to and from alternate hosts. National Meeting of the Entomological Society of America. 10 minute oral presentation. November 2021.
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Strayer-Scherer, A.; Hagan, A. K.; Burch, K.; Conner, K., Brown, S. M.; Koebernick, J.; Zaccaron, M.; Miller, H. B.; and Canterbury, J. 2022. Cotton leafroll dwarf virus incidence as impacted by planting date and cotton cultivar in Alabama. During Proceedings of the Beltwide Cotton Conferences (San Antonio, TX).
• Type: Conference Papers and Presentations Status: Published Year Published: 2022 Citation: Adegbola, R. O.; Kitchen, N. T.; Lawrence, K.; Conner, K.; Mulvaney, M. J.; Small, I.; Price, P.; Kerns, D. L.; Huseth, A. S.; Strayer-Scherer, A.; Zaccaron, M.; Greene, J.; Ali, A.; and Brown, J. K. 2022. Surveillance of Cotton leafroll dwarf virus (CLRDV) in the US by molecular diagnostics and phylogeny of ORF0 and ORF3 fragments. During Proceedings of the Beltwide Cotton Conferences (San Antonio, TX).

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

Outputs
Target Audience:The target audiences of these projects include the research community, university extension entomologists, county and regional extension agents, crop consultants and producers in the Southern U.S. who are managing cotton. Changes/Problems:No changes have been made to the approach but the ongoing pandemic continues to cause delays in supply chains, shortages in consumables needed for molecular biology protocols, and delays in sequencing services. Many meeting venues have used online formats, and many local (state) meetings have been canceled since the start of the pandemic. All planned international conferences and trips have been cancelled due to COVID travel restrictions and safety concerns. Despite these challenges we are continuing to adapt and make progress on experiments and dissemination of results. A no cost extension was received to extend the end date of the project to August 31, 2022. What opportunities for training and professional development has the project provided?Co-PIs, Students and postdocs have presented research related to CLRDVat virtual national society meetings during the past fiscal year. PI and co-PI participation in regional working group meetings has enabled us to begin coordinating regional research efforts in entomology, plant pathology and plant breeding to expand research efforts beyond the scope of this grant and get similar data on symptomatology, aphid dispersal, and varietal differences in susceptibility across the southeast and mid-south. How have the results been disseminated to communities of interest?The current state of knowledge about CLRDV distribution, incidence, spread and management have been disseminated at local and regional extension meetings, annual working group meetings, and annual meetings of professional societies (including, Entomological Society of America and Annual Phytopathological Society, the Beltwide Cotton Conferences, and the National Conservation Systems Cotton and Rice Conference). Preliminary results along with diagnostic guides with images have been disseminated to stakeholders via articles in print and electronic media, and social media outlets. Information shared at these venues and in extension publications has been disseminated by several popular media outlets, including Farm Press and local media. What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period data collection will finish in the fall of 2021. Research analyses will be conducted and disseminated at annual meetings during 2022. Scientific peer-reviewed manuscripts will be submitted for publication and extension outlets will be used to disseminate updates throughout the next fiscal year.

Impacts
What was accomplished under these goals? Cotton leafroll dwarf virus (CLRDV) is an emerging aphid transmitted virus in the US cotton belt. CLRDV was first identified in cotton samples collected in AL in 2017 and has been confirmed in cotton growing states from VA to west TX. Yield impacts to-date have been difficult to quantify, but fields with 100% yield loss have occurred at several locations in GA, AL and MS. Fundamental knowledge of the epidemiology and efficacy of management practices for reducing virus incidence and yield loss are needed to mitigate economic losses caused by this virus. Understanding insect dispersal events that result in colonization of cotton and virus transmission is a cornerstone of developing sound management practices for insect transmitted viruses. This information will help identify which disease and aphid management practices to include in an integrated management plan and inform how and when they should be used. This multi-state, multi-disciplinary effort between Auburn University (AU) and the University of Georgia (UGA) aims to identify management practices that can be used by growers, and to disseminate information about the spread and management of CLRDV to research and stakeholder communities. Objectives Objective 1. Characterize aphid population dynamics in cotton agroecosystems and identify the timing of primary spread of CLRDV to cotton (Research Objective). Trapping programs to monitor aphid flights were initiated in March 2021 at two locations in GA and at three locations in AL. Aphids captured in traps are collected weekly and stored in ethanol until they are identified. Trapping efforts are ongoing, and weekly trapping will continue through the end of the growing season (October-November 2021). Aphid counting, identification and data recording of AL and GA trap samples is being conducted at AU. Weekly monitoring of virus spread using sentinel plants was initiated in April and is conducted concurrent with aphid trapping at the three locations. Sentinel plant monitoring in 2021 is planned until the end of November. Virus testing of sentinel plant cohorts is ongoing and will conclude 2.5 month after the last cohort of plants is collected from the field. Objective 2. Evaluate the efficacy of host plant resistance, vector management and cultural practices on reducing CBD incidence, disease severity and yield loss (Research & Extension Objective). Objective 2a (Entomology): Field trials examining the effects of aphid management and planting date have been completed at AU (PI, Jacobson) and UGA (co-PIs Roberts and Toews). These field trials provided conclusive data that making foliar applications of insecticides for aphid management, including intensive weekly sprays, were not effective at reducing the final incidence of CLRDV. Virus incidence ranged from 60-100% and 90-100% per plot in AL and GA, respectively. These results are valuable because they communicate that aphid management in high-pressure areas is not an effective method for reducing virus transmission. This information is being disseminated to stakeholders to prevent an increase in insecticide use. A manuscript of the results is currently in review. Jacobson, Roberts and Toews are continuing a small-scale version of this experiment to generate updated information to share with growers to reduce unnecessary increases in insecticide use targeting aphids. These trials are conducted with two planting dates, non-sprayed control plots, and a weekly-spray treatment. Aphids were counted weekly in these plots. Insecticide treatments have terminated. Cotton samples will be collected and tested for CLRDV in September 2021. Objective 2b (Plant breeding): The combined efforts of field- and molecular-based plant breeding research programs at AU and UGA are identifying genetic and phenotypic characteristics of resistance that will identify breeding strategies for developing elite germplasm that is resistant to disease losses caused by CLRDV. At UGA (Chee), marker trait association studies showed promising results that identified putative resistance genes in two populations that segregated for viral symptoms. The DNA of these two populations were sampled and sent to Texas A&M Genome Center for additional sequencing to confirm results from the analyses conducted at UGA. Sequencing data arrived late-August 2021 and will be analyzed this fall. At AU (Koebernick), 1,200 cotton varieties were phenotyped for CLRDV symptoms and infection status in experiments conducted at two locations in 2019 and 2020. There were 5 lines identified as being negative for the virus during both field seasons. Another replicate of these experiments is being conducted in 2021 with fewer lines, including the 5 that tested negative for CLRDV in 2019 and 2020. Samples will be collected and CLRDV infection status will be determined in September of 2021. Varieties that tested negative from these trials will be tested for infection in laboratory experiments that use a leaf-disc assay. These assays are reflective of a local infection immediately (3-5 days) after transmission. If leaf discs become infected it would suggest that a virus movement blocking mechanism may be responsible for observed resistance. The results of these assays will inform future experiments to better understand the probability of systemic infection and whether or not host plant defenses can suppress subsequent viral replication, movement and disease expression in these lines. Objective 3. Develop educational materials and programs to increase stakeholder knowledge and adoption of an integrated pest management program for managing CBD (Extension Objective). Final data analysis from research efforts completed in the fall of 2020 have provided information that has been, and continues to be, shared with stakeholder communities of interest. Updated extension publications on CLRDV were created and disseminated at both UGA and AU. As a Center of Excellence for CLRDV research in the U.S. we have also worked to secure funding to help extend our research networks and develop new collaborations with research and extension personnel at land grant universities across the southeast and mid-south. Drs. Jacobson, Bowen, Conner, Hagan, and Koebernick are collaborators on a recently funded Southern IPM Center Working Group grant lead by Dr. Amanda Strayer-Scherer, a cotton extension pathologist hired at AU in 2019. This grant is supporting regional sentinel plots to examine variety x environment x CLRDV interactions, virus incidence and symptomatology. Collaborators include plant pathologists, entomologists, breeders and agronomists from VA, NC, SC, GA, FL, MS, TN, LA, AK, and TX in 2020-present. Drs. Conner and Bag also continue to provide support for diagnostics needed by land grant extension and research personnel in neighboring states to test suspect fields.

Publications

• Type: Journal Articles Status: Published Year Published: 2021 Citation: Tabassum, A., Bag S., Suassuna, N. D., Conner, K. N., Chee, P., Kemerait, R.C., and Roberts, P. (2021). Genome analysis of cotton leafroll dwarf virus reveals variability in the silencing suppressor protein, genotypes, and genomic recombinants in the USA. PLoS ONE. 16(7): e0252523. https://doi:10.1371/journal.pone.0252523
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Sedhain, N. P., Bag, S., Carter, R., Morgan, K., Triana, P., Kemerait, R. C., Roberts, P. M. (2021). Natural host range, incidence on overwintering cotton and diversity of cotton leafroll dwarf virus in Georgia USA. Crop Protection. 144:105604. https://doi.org/10.1016/j.cropro.2021.105604
• Type: Journal Articles Status: Published Year Published: 2021 Citation: Bag, S., Roberts P. M., and Kemerait R.C. (2021). Cotton leafroll dwarf virus: an emerging virus disease on cotton in the US. Crops and Soils 54:2, 18-22. (Feature Article). https://doi.org/10.1002/crso.20105
• Type: Journal Articles Status: Submitted Year Published: 2022 Citation: Mahas, J., P.M. Roberts, K. Conner, S. Bag, E. K. Blythe, M. Toews, and A.L. Jacobson. Planting date and Aphis gossypii management on final incidence of Cotton leafroll dwarf virus. Crop Protection.
• Type: Journal Articles Status: Under Review Year Published: 2022 Citation: Conner, K.; Sikora, E.; Koebernick, K.; and Zaccaron, M. 2021. Interdisciplinary Team Addresses Cotton Leafroll Dwarf Virus in Alabama. Journal of Extension. In Review.
• Type: Journal Articles Status: Under Review Year Published: 2022 Citation: B. Heilsnis, J. Koebernick, A. Jacobson, and K. Conner. 2020. Investigating the Vector Competency of Aphis gossypii and Bemisia tabaci MEAM1 to Transmit Cotton leafroll dwarf virus. Phytofrontiers. In review.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Roberts, P., M. Toews, S. Taylor, D. Reisig, A. Huseth, F. Reay-Jones J.K. Greene, S.V. Paula-Moraes, A. Jacobson, S. Graham, and R. Smith. 2021. Cotton aphid insecticide efficacy in the southeast: a two-year summary. National Cotton Council Beltwide Cotton Conferences.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Mahas, J.W., P.M. Roberts, K. Conner, S. Bag, M.D. Toews, E.K. Blythe, F.B. Hamilton, C.H. Ray, A.L. Jacobson. November 2020. Managing the Primary vector of Cotton leafroll dwarf virus does not Reduce Final Virus Incidence in Cotton. The Annual Meeting of the Entomological Society of America Virtual Annual Meeting. Poster Presentation. Second place graduate student research poster for PIE section.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Mahas, J.W., A.L. Jacobson. October 2020. Quantifying Imidacloprid Susceptibility in Populations of Aphis gossypii. Auburn University's Graduate Student Poster Showcase. Auburn, AL. Virtual Conference. Poster presentation. First place graduate student research poster.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Mahas, J.W. and A.L. Jacobson. October 2020. Quantifying Imidacloprid Susceptibility in Populations of Aphis gossypii. Southeastern Population Ecology & Evolutionary Genetics 2020 Virtual Conference. Poster presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Jacobson, A.L., C. Ray, P. Roberts, M. Toews, J. Greene, A.S. Huseth, G.G. Kennedy, S. Taylor, S. Paula-Moraes, S. Brown, J. Gore, W. Crow, S.D. Stewart, D. Kerns, S. Vyavhare, S.H. Graham, R. Smith and K. Kesheimer. January 2021. Monitoring Population Dynamics of Aphids across the Cotton Belt. Proceedings of the Beltwide Cotton Conferences. Virtual Conference. 15 minute oral presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Zaccaron, M., J. Greene, A.K. Hagan, A. Strayer-Scherer, T.W. Allen, J. Koebernick, A.L. Jacobson, K.L. Bowen, T.R. Faske, P. Price, I. Small, M.J. Mulvaney, R. Kemerait, A.S. Huseth, H.M. Kelly, D. Kerns, S. Taylor and H. Wang. January 2021. Incidence and Impact of Cotton Leaf Roll Dwarf Virus (CLRDV) in the USA Cotton Belt. Proceedings of the Beltwide Cotton Conferences. Virtual Conference. 15 minute oral presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Heilsnis, B., J. Koebernick, K. Conner, A.L. Jacobson. January 2021. Rapid Screening for Resistance to CLRDV: Development of an Aphid Assay. Proceedings of the Beltwide Cotton Conferences. Virtual Conference. 15 minute oral presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Mahas, J.W., A. S. Huseth, A.L. Jacobson. January 2021. Quantifying Imidacloprid Susceptibility in Alabama Populations of Aphis gossypii. Proceedings of the Beltwide Cotton Conferences. Virtual Conference. 15 minute oral student presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Roberts, P., M. Toews, S. Taylor, D. Reisig, A.S. Huseth, F. Reay-Jones, J. Greene, S. Paula-Moraes, A.L. Jacobson, S. Graham and R. Smith. January 2021. Cotton Aphid Insecticide Efficacy in the Southeast: A Two Year Summary. Proceedings of the Beltwide Cotton Conferences. Virtual Conference. 15 minute oral student presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: A. K. Hagan, A. Strayer-Scherer, K. Burch, K. Bowen, H.B. Miller, C. McElmoyl, and J. Burkett. 2021. Assessment of Yield loss Attributed to Cotton leafroll Dwarf Disease in Alabama Cotton. Beltwide Cotton Conferences Proceedings 2021.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: A. Strayer-Scherer, A. K. Hagan, K. Burch, K. Conner, S. Brown, J. Koebernick, M. Zaccaron, H. B. Miller, and D. Moore. 2021. Occurrence of Cotton Leafroll Dwarf Disease as Impacted by Cotton Cultivar and Planting Date. Beltwide Cotton Conferences Proceedings 2021.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Invited Presentation: Jacobson, A.L., and P. Roberts. February 2021. Update on Aphid Vectors, Virus Spread and Management of Cotton Leafroll Dwarf Virus. National Conservation Systems Cotton & Rice Conference. Online. 50 minute oral presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Invited Presentation: Jacobson, A.L. December 2020. Update on Aphid Vectors and Management of Cotton Leafroll Dwarf Virus. 32th Annual Texas Plant Protection Conference. Online. 15 minute oral presentation.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Invited Presentation: Roberts, P. 2021. Management of cotton aphid and implications for CLRDV. 2021 Cotton and Rice Conference.
• Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Roberts, P., M. Toews, S. Taylor, D. Reisig, A. Huseth, F. Reay-Jones, J. Greene, S. Paula-Mores, A. Jacobson, S. Graham, and R. Smith. 2021. Grow Plant Health Exchange Webinar found at: https://www.planthealthexchange.org/cotton/Pages/GROW-COT-06-21-309.aspx
• Type: Other Status: Published Year Published: 2021 Citation: Roberts, P., and M. Toews. (1997-present, annual update). Cotton insect control. In: Georgia Pest Control Handbook Commercial Edition, (P. Guillebeau, Ed.). The Univ. of Georgia Coop. Ext. Ser. Special Bull. 28. Found online at http://www.ent.uga.edu/pmh.
• Type: Other Status: Published Year Published: 2021 Citation: Conner, K.N., A. Strayer-Scherer, A. Hagan, J. Koebernick, A.L. Jacobson, K. L. Bowen, E. Sikora, S. Graham, and S. Brown. 2021. Cotton Leafroll Dwarf Virus. ANR-2539 https://www.aces.edu/blog/topics/crop-production/cotton-leafroll-dwarf-virus/
• Type: Other Status: Published Year Published: 2021 Citation: Selection of Popular Press Articles that wrote stories after hearing information at meetings: https://www.farmprogress.com/cotton/cotton-virus-prevalent-sky-isnt-falling https://alabamanewscenter.com/2020/10/18/auburn-university-research-team-tackles-new-cotton-virus/ https://www.dtnpf.com/agriculture/web/ag/crops/article/2021/01/26/cotton-leafroll-dwarf-virus-poses-us https://agfax.com/2020/02/21/alabama-cotton-leaf-roll-dwarf-virus-what-you-should-know/ https://extension.uga.edu/story/8708/Cotton-Leaf-Roll-Dwarf-Virus.html https://www.dailyadvent.com/news/amp/1a699a24cfda60ec93f8d84cb10c6192-Cotton-virus-confirmed-in-24-Georgia-counties https://www.wrganews.com/2021/08/23/cotton-virus-found-in-24-georgia-counties/ https://www.wrganews.com/2021/08/23/cotton-virus-found-in-24-georgia-counties/ https://www.newsbreak.com/news/2348138096331/cotton-virus-confirmed-in-24-georgia-counties http://ocm.auburn.edu/newsroom/news_articles/2020/10/080920-cotton-virus-research.php

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

Outputs
Target Audience:The target audiences of these projects include the research community, university extension entomologists, county and regional extension agents, crop consultants and producers in the Southern U.S. who are managing cotton. Changes/Problems:Major disruptions in research operations were experiences at AU and UGA due to the emerging COVID-19 pandemic. COVID restrictions also slowed accounts being set-up at AU and UGA because all support staff worked from home fromfrom March-August and communication with our Contracts and Grants departments within and across universities were limited. At AU, ongoing research was classified as "essential', and therefore progress on this project continuedunder strict guidelines that limited hiring, the number of personnelcleared for travel and number of people working in the same lab/greenhouses. This changed our operations and the volume of research we normally conduct. We continue to work under restrictions, butAU'sCOVID policies have not been as strict as our collaborators in Georgia.The state of Georgia enacted a "shelter in place" mandate starting April 3 and lasting until May 1. No one was allowed to come into the office or conduct any lab or field work (other than maintaining living organisms) during this period. Further, UGA enacted a tiered approach to allowing faculty to conduct research that started June 1. We missed much of the early spring planning window and were not allowed to collect data during much of the early summer. This action severely hampered our ability to collect time sensitive data. Further, we continue to be under an out of state travel ban and are not allowed to attend professional conferences. Due to COVID-related restrictions, the field experiments conducted under the entomology objectives have been modified based on our current work capabilities. The two-site years of data collected on the aphid management trial included in the original proposal were in total agreement that foliar applications were not effective at reducing CLRDV transmission (results will be submitted for publication this fall). During this trial wenot able to assess symptomatology or effects of planting date (plant age)because the disease pressure wasvery high across all plots (60-100% infected plants in all plots). In AL we are continuing research to examinerelationships between thetiming of inoculation (plant age) and yield loss by coveringour field plots with cages covered with aphid-proof screen (2 rows wide x 20' long), and releasing viruliferous aphids in them at prescribed times. This allows us tocontrol the timing of inoculation at specific crop growth stages and examine timing of infection x yield loss interactions among four release dates. Weekly releases of viruliferous aphids were performed from emergence to the7-9 true-leaf growth stage in 2020. In 2019 the first year of data collection was performed for this experiment using funds from a one-year FFAR grant. The 2019 trials showed a significant reduction in yield compared to the last release date and controls (no-aphid and non-viruliferous aphid treatments). Although not significant, there was an incremental trend for a numerically lower yield reduction in the second and third release datesthat wasgreater in the second. Due to the success in using this experimental design last year, CPPM funds will be used to continue this study in 2020 and 2021. Data collected from this trial includes the proportion of plants infested with aphids after release, the average number of aphids per plant, plant mapping data to examine symptoms and boll retention at the first and second fruiting branch positions (responsible for majority of yield), yield, and fiber quality. Disruptions at UGA limited the number of trials they were able to conduct, but three insecticide efficacy trials were conducted in which they collected additional data on aphid populations dynamics in the crop, population reductions after management, and CLRDV symptoms. The large (15 treatment) planned trial reached threshold in early July and was sprayed on time; we scouted in before and after the application and continue to monitor this trial. The second trial was planted in Bleckley County on a commercial farm with 6 currently recommended insecticides. This trial also triggered and we counted aphids before and after the treatments and continue to monitor. The third trial was planted on May 10, 2020 using ST5122 cotton, which is susceptible to cotton blue disease. Plots were monitored weekly for cotton aphids when populations started building in mid-June. On July 15, after four weekly sampling events, aphid fungus was detected in the field before cotton aphid populations reached the treatment threshold. Therefore, this field did not get sprayed this year. We are maintaining the field for use in whitefly or other late season cotton insect pests. The UGA microginhas, and is continuing to gin cotton lint samples collected from all field trials conducted at AU and UGA. Entomologists at AU and UGA will coordinate 2021 trials to continue to examine the interaction between cultural management practices such as planting date, and disease severity. The design and scope of the trials will be dependent upon the status of the pandemic and research restrictions in place. Breeding efforts and plant pathology objectives have remained the same and progress are continuing as planned. What opportunities for training and professional development has the project provided?Co-PIs, Students and postdocs have presented research related to CLRDVat national society meetings during the fall of 2019 andprior to COVID shutdowns that beganMarch 2020. PI and co-PI participation in regional working group meetings has enabled us to begin coordinating regional research efforts in entomology, plant pathology and plant breeding to expand research efforts beyond the scope of this grant and get similar data on symptomoatology, aphid dispersal, and varietal differences in susceptibility across the southeast and mid-south. How have the results been disseminated to communities of interest?The current state of knowledge about CLRDV distribution, incidence, spread and management have been disseminated at local and regional extension meetings (including the Mississippi Row Crops Short Course, Georgia Farm Bureau annual conference, UGA Tifton Campus Cotton and Peanut Research Field Day), annual working group meetings (including, Cotton Inc. Research Review and Planning Meeting on Cotton Leafroll Dwarf Virus, Southern Pest Management Seminar and Southeastern Row Crop Entomology Working Group), and annual meetings of professional societies (including, Entomological Society of America and Annual Phytopathological Society, the Beltwide Cotton Conferences, and the National Conservation Systems Cotton and Rice Conference). Information was also disseminated to international cotton research communities at the AACS 2019 Australian Cotton Research Conference in Australia, where management programs have been developed for a closely related cotton-infecting polerovirus, Cotton bunchy top virus. Other regional, national and international conferences and trips were planned in 2020, but cancelled due to COVID travel restrictions and safety concerns. Research Presentations at scientific meetings: Mahas, J. and A. Jacobson. November 2019. Susceptibility of Aphis gossypii to imidacloprid in Alabama, USA. The Annual Meeting of the Entomological Society of America. St. Louis, MO. Poster presentation. First place graduate student research poster for PIE section. Brown, S., K. Conner, A. Hagan, A. Jacobson, J. Koebernick, K. Lawrence, S. Bag, B. Kemerait, P. Chee, T. Allen, S. Sabanadzovic, and B. Nichols. 2019. Report of a Research Review and Planning Meeting on Cotton Leafroll Dwarf Virus. October 8, 2019. Orange Beach, AL. https://www.cottoninc.com/wp-content/uploads/2019/11/10-19-CLRDV-Research-Review-Meeting-Report-Nichols.pdf Chee, P. Cotton (new) disease: symptoms development and inheritance. Cotton Leafroll dwarf virus research review meeting. Orange Beach, AL. October 8, 2019. Peng Chee Addressing Cotton Blue Disease in the USA. In AACS 2019 Australian cotton research conference. UNE Armidale, Australia. October 29, 2019. Bag, S. 2019. Cotton leafroll dwarf disease: an emerging threat to cotton production in GA. In: Georgia Farm Bureau annual conference, Jekyll Island, 12/09/2019 Jacobson, A.L., P. Roberts, M. Toews, K. Conner, S. Bag, J. Mahas, A. Kesheimer. 2020. Investigating the Impact of Planting Date and Cotton Aphid Management on the Final Incidence of Cotton Leafroll Dwarf Virus. Proceedings of the Beltwide Cotton Conferences, Austin, TX. 15 minute oral presentation. Heilsnis, B., K. Conner, J. Koebernick, Jacobson, A.L. 2020. Transmission of Cotton Leafroll Dwarf Virus By Aphis Gossypii. Proceedings of the Beltwide Cotton Conferences, Austin, TX. 15 minute oral presentation. Roberts, P., M. Toews, A.L. Jacobson, R. Smith, S. Paula-Moraes, J. Greene, D. Reisig, S. Taylor. 2020. Cotton Aphid Insecticide Efficacy in the Southeast. Proceedings of the Beltwide Cotton Conferences, Austin, TX. 15 minute oral presentation. McLaughlin, A., K. Conner, K.L. Bowen, A.K. Hagan, W. Groover, K. Lawrence, and A.L. Jacobson. 2020. Leafroll Dwarf Virus Inoculation on Disease Severity and Yield Loss. Proceedings of the Beltwide Cotton Conferences, Austin, TX. 15 minute oral presentation. Lawrence, K., A. Hagan, R. Norton, J. Hu, T. Faske, R. Hutmacher, J. Mueller, I. Small, Z. Grabau, B. Kemerait, D. Jardine, P. Price, T. Allen, C. Meeks, J. Idowu, L. Thiessen, S. Byrd, J. Goodson, H. Kelly, T. Wheeler, T. Isakeit, and H. Mehl. 2020. Cotton Disease Loss Estimate Committee Report 2019. Proceedings of the Beltwide Cotton Conferences, Austin, TX. oral presentation Zaccaron, M., A. Hagan, K. Conner, and J. Koebernick. 2020. CLRDV multi-state sentinel plot and screening trial results. Proceedings of the Beltwide Cotton Conferences, Austin, TX. 15 minute oral presentation. Koebernick, J. 2020. Addressing cotton blue disease detected in the US. National Conservation Systems Cotton and Rice Conference. January 30-31, Memphis, TN. oral presentation Bag, S. 2020. Aphid-transmitted Cotton leafroll dwarf disease: An emerging threat to sustainable production of cotton. In: APS annual meeting, August 8-12, 2020. Extension Publications Bag, S. and P. Roberts. 2019. Cotton Leafroll Dwarf Disease: An emerging threat to cotton production in GA. 82nd Annual GFB Convention Roberts, P. 2019. Cotton Insect Management Update. 82nd Annual GFB Convention. Roberts, P. 2019. Aphid and SLWF Management. UGA Tifton Campus Cotton and Peanut Research Field Day. Jacobson, A.L., K. Conner, and P. Roberts. December 2019. CLRDV - What We've Learned from an Entomological Perspective. Mississippi Row Crops Short Course. Starkville, MS. 30 minute oral presentation. Roberts, P. and A.L. Jacobson. December 2019. Aphid Management and Implications for CLRDV. Mississippi Row Crops Short Course. Starkville, MS. 30 minute oral presentation. Jacobson, A.L., P. Roberts, and M. Toews. October 2019. The Aphid Vector. Cotton Leaf Roll Dwarf - like Virus (CLRDV) Research Review and Planning Meeting. Gulf Shores, AL. 15 minute oral presentation. Roberts, P., A.L. Jacobson, and M. Toews. October 2019. Aphid Management. Cotton Leaf Roll Dwarf - like Virus (CLRDV) Research Review and Planning Meeting. Gulf Shores, AL. 15 minute oral presentation. Hagan, A. Twenty-four Twitter posts related to CLRDV (2018-2020). Roberts, P. 2020. GA County Cotton Production Meetings. In total 30 meetings were conducted with 1,569 attendees. CLRDV and aphid management were included in all programs. What do you plan to do during the next reporting period to accomplish the goals?Data collection and research conducted to monitoring aphid population dynamics, CLRDV spread, and cotton breeding will continue in 2021. Results of research conducted in 2019 and 2020 will be used to generate scientific manuscripts and extension communications that will bedisseminated to research and stakeholder communities throughout the next fiscal year.

Impacts
What was accomplished under these goals? Cotton leafroll dwarf virus (CLRDV) is an emerging aphid transmitted virus in the US cotton belt. CLRDV was first identified in cotton samples collected in AL in 2017 and has been confirmed in cotton growing states from VA to west TX. Yield impacts to-date have been difficult to quantify, but fields with 100% yield loss have occurred at several locations in GA, AL and MS. Fundamental knowledge of the epidemiology and efficacy of management practices for reducing virus incidence and yield loss are needed to mitigate economic losses caused by this virus. Understanding insect dispersal events that result in colonization of cotton occur, and when these events contribute to spread of plant viruses from the non-crop reservoir species to crop species is a cornerstone of developing sound management practices for insect transmitted viruses. This information will help identify which disease and aphid management practices should comprise an integrated management plan and inform how and when they should be used. This multi-state, multi-disciplinary effort between Auburn University (AU) and the University of Georgia (UGA) aims to identify management practices that can be used by growers, and disseminate information about the spread and management of CLRDV to research and stakeholder communities. Objectives Objective 1. Characterize aphid population dynamics in cotton agroecosystems and identify the timing of primary spread of CLRDV to cotton (Research Objective). Trapping programs to monitor aphid flights were initiated in late-March at two locations in GA and the first week of March at three locations in AL. Aphids captured in traps are collected weekly and stored in ethanol until they are identified. Trapping efforts are ongoing, and weekly trapping throughout the grant period is planned. Aphid identification and data recording of AL and GA trapping samples is being conducted at AU. Weekly monitoring of virus spread using sentinel plants was initiated in April in AL and is conducted concurrent with aphid trapping at the three locations. Sentinel plant monitoring in 2020 is planned until the end of November, and will be repeated again in 2021. Virus testing of sentinel plant cohorts at AU is ongoing. Sentinel plant monitoring in 2020 was not possible at UGA due to complications getting clearance to conduct the greenhouse and field research during the statewide COVID-19 shelter in place mandate. UGA VP for Research David Lee refused to allow PIs to start new projects, only to continue projects that included data from previous years. Ongoing sentinel plant monitoring in both states is planned for 2021. Objective 2. Evaluate the efficacy of host plant resistance, vector management and cultural practices on reducing CBD incidence, disease severity and yield loss (Research & Extension Objective). Objective 2a (Entomology): Field trials examining the effects of aphid management and planting date have been completed at AU (PI, Jacobson) and UGA (co-PIs Roberts and Toews). These field trials provided conclusive data that making foliar applications of insecticides for aphid management, including intensive weekly sprays, were not effective at reducing the final incidence of CLRDV. Virus incidence ranged from 60-100% and 90-100% per plot in AL and GA, respectively. These results are valuable because they communicate that aphid management in high-pressure areas is not an effective method for reducing virus transmission. This information is being disseminated to stakeholders to prevent an increase in insecticide use. A manuscript of the results is currently in preparation and will be submitted for review this fall. Objective 2b (Plant breeding): The combined efforts of field- and molecular-based plant breeding research programs at AU and UGA are identifying genetic and phenotypic characteristics of resistance that will identify breeding strategies for developing elite germplasm that is resistant to disease losses caused by CLRDV. In 2019, UGA (co-PI, Chee) tested the hypothesis that the Brazilian line BRS286 which is resistance to Blue disease in Brazil, also is resistant to the stain of CLRDV in the US. Two genetic populations were developed using BRS286 as the resistant parent and tested for segregation of resistance in a disease nursery near Fairhope, Alabama (co-PI, Koebernick). Unfortunately, BRS286 was determined to be susceptible to the CLRDV in Alabama and Georgia. Interestingly, several F2 populations in our breeding nursery were observed as segregating for the viral symptoms, particularly those crosses that involved BRS286. Therefore, in the first quarter of 2020, major efforts at UGA focused on extracting DNA from two F2 populations derived from the BRS286 parent. The DNA samples were sent to the Texas A&M Genome Center on May 20th for genotyping using the high-throughput 63K SNP array. The genotyping was delayed due to COVID-19 and we received the data on July 24th. Marker trait association analysis is currently in progress at UGA. Experiments were conducted at two locations in AL (Koebernick) in 2019 to screen 1,200 varieties for their susceptibility to CLRDV under field conditions. In these experiments6% of the varieties tested negative for CLRDV using PCR-based methods at the end of the season. In 2020, these lines were tested for resistance using a greenhouse and laboratory-based leaf-disk assay to determine whether CLRDV replication can be detected in leaf tissue after aphid transmission. Results indicate that these lines can be infected with the virus, but these assays are reflective of a local infection immediately (3-5 days) after transmission. Future research is needed to better understand the probability of systemic infection and whether or not host plant defenses can suppress subsequent viral replication, movement and disease expression in these lines. In 2020 an additional 600 lines have been planted at the plant breeding unit in Tallassee, AL, and 400 of these lines are duplicated in Fairhope, AL. Symptoms are being observed season long and each plot will be subjected to PCR testing to confirm incidence. Objective 3. Develop educational materials and programs to increase stakeholder knowledge and adoption of an integrated pest management program for managing CBD (Extension Objective). Final data analysis from research efforts completed in the fall of 2020 will provide information needed to begin work on this objective. As a Center of Excellence for CLRDV research in the U.S. we have also worked to secure funding to help extend our communication networks and develop new collaborations with research and extension personnel at land grant universities across the southeast and mid-south. Drs. Jacobson, Bowen, Conner, Hagan, and Koebernick are collaborators on a recently funded Southern IPM Center Working Group grant lead by Dr. Steve Brown, a cotton extension agronomist hired at AU in 2019, to organize an annual meeting between plant pathologists, entomologists, breeders and agronomists from VA, NC, SC, GA, FL, MS, TN, LA, AK, and TX in 2020 and 2021. In-person meetings during the fall of 2020 have been cancelled due to COVID-related travel restrictions and state mandates.

Publications

• Type: Journal Articles Status: Awaiting Publication Year Published: 2020 Citation: In preparation, to be submitted fall 2020, USDA funding acknowledged: Mahas, J., P.M. Roberts, K. Conner, S. Bag, E. K. Blythe, M. Toews, and A.L. Jacobson. Fall 2020 submission date. Planting date and Aphis gossyppii management on final incidence of Cotton leafroll dwarf virus. Targeted journal: Pest Management Science.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Wang, H., J. Greene, J.D. Muller, K. Conner and A. Jacobson 2020. First report of Cotton leafroll dwarf virus in cotton fields of South Carolina. Plant Disease. https://doi.org/10.1094/PDIS-03-20-0635-PDN Funded with matching fund portion of this project from Cotton Inc.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Thiessen, L. T.L. Schappe, M. Zaccaron, K. Conner, J. Koebernick, A. Jacobson, and A. Huseth. 2020. First report of cotton leafroll dwarf disease caused by Cotton leafroll dwarf virus affecting cotton in North Carolina. Plant Disease. https://doi.org/10.1094/PDIS-02-20-0335-PDN Funded with matching fund portion of this project from Cotton Inc.
• Type: Journal Articles Status: Published Year Published: 2020 Citation: Tabassum, A., P.M. Roberts, and S. Bag. (2020). Genome Sequence of Cotton Leafroll Dwarf Virus Infecting Cotton in Georgia, USA.. Microbiol Resour Announc, 9(34). doi:10.1128/MRA.00812-20. Funded with matching fund portion of this project from Cotton Inc.