Recipient Organization
UNIVERSITY OF GEORGIA
200 D.W. BROOKS DR
ATHENS,GA 30602-5016
Performing Department
(N/A)
Non Technical Summary
We propose to develop a dynamic risk model and climate suitability model based on existing climate data and current observational/experimental data to be generated through these research objectives: (1) document the distribution of HLB and ACP in cold-hardy citrus growing regions of south Georgia, north Florida and east-central California, (2) evaluate preference of ACP to cold-hardy citrus cultivars, (3) determine the titer of CLas in trees following exposure to cold and freezing events, and (4) tolerance and acclimatization in ACP populations to cold and determine underlying mechanisms in the insect's body to these adaptations. The proposal addresses three priority areas stated by the Citrus Disease Subcommittee (CDS) and ECDRE program: (#2) regional management or eradication of ACP, (#3) predictive models of psyllids movement and dispersal, early detection of HLB, and (#9) greater understanding of the ecology and interactions of the citrus production system and HLB disease complex. Citrus production acreage in areas with cool winter temperatures is increasing and the sustainability of cold-hardy citrus production in parts of California, Florida and Georgia. The lack of scientific studies on the effect of cold on both CLas and ACP populations leaves cold-hardy citrus production in US at risk and must be addressed. Our proposed objectives will provide basic information on the influence of cold on vector and pathogen biology, which would help in developing region-specific risk models to help in decision-making process for effective sampling, surveillance and future expansion of groves.
Animal Health Component
45%
Research Effort Categories
Basic
55%
Applied
45%
Developmental
0%
Goals / Objectives
The main goal of this project is to assessthe risk of HLB in cold-hardy citrus production regions and develop modeling tools to help in decision-making for vector and pathogen surveillance and identifying "low-risk" areas for future citrus expansions. In addition, the impact of cold on ACP and CLas in cold-hardy citrus production areas will be determined. This information is fundamental to establishing whether HLB poses as great a threat to cold-hardy production areas as it has in more subtropical areas. This will be important for growers considering moving citrus production northward. The current distribution of ACP and HLB will be established, and relationships with cold temperatures typical of cold-hardy production areas will be evaluated. The potential for cold adaptation of ACP will also be determined. Information will be generated on the preference of ACP to common cold-hardy citrus cultivars.Finally, a better understanding of how cold and freeze events impact the spread and survival of theCLas bacterium that causes HLB and ACP will be particularly important for determining HLB risk to the cold-hardy citrus industries and for developing appropriate management strategies in cold-hardy citrus production areas.
Project Methods
Obj. 1aProducing Citrus Layer in north FL and south GA:To generate an accurate citrus layer, we will acquire remote sensing data from CropScape datasets provided by NASS for the targeted regions. The extracted layers will focus on citrus-related land use classifications. Concurrently, we will obtain detailed commercial citrus location data. In the identification phase, we will use GIS analysis to pinpoint potential citrus groves by analyzing the CropScape data and applying filters to isolate areas designated as citrus classification. We will conduct geospatial analysis to map citrus areas in north FL and south GA. The final citrus layer will be exported in formats like shapefiles and GeoJSON for easy integration into GIS platforms and analysis.Obj. 1bSampling for HLB, ACP monitoring andCLas detection: We will collect leaf samples forCLas detection and establish a monitoring plan for ACP presence in selected locations in south GA and north FL. Symptomatic trees will be prioritized for leaf sampling collection, but asymptomatic trees will also be included to detect early-stage infections. Leaf samples will be collected in spring and fall. Sampling and monitoring for ACP will also be conducted twice each year. Both leaf and insect samples will be subjected to molecular detection forCLas bacteria using a real-time PCR assay.Obj. 1c Outreach and Extension:To gain the stakeholders input on potential risk of HLB occurrence in cold-hardy citrus, team members will engage and hold Extension meetings, and field days. Program evaluation will be carried out twice a year: in spring and fall meetings. Based on the feed-back and evaluation results, we will improve our outreach program to make more impactful.Obj. 2aLife-table studies: Laboratory experiments will estimate ACP population growth rates on common cold-hardy citrus. Five cultivars will be evaluated: two mandarins that are commonly grown in colder regions in FL, GA, and CA.To initiate the life-table studies, 20 colony-sourced,CLas-free ACP adults will be caged on citrus plants. After three days, adults will be removed, and eggs counted using a 10x hand lens. Plants will be examined every other day using a 10x hand lens and the number of ACP by life stage (eggs, first-third instars, fourth-fifth instars, and adults) will be recorded. Counts will continue until all immature ACP emerge as adults or die. Survivorship and development will be used to construct life tables for each citrus cultivar according to Liu and Tsai (2000).Obj. 2bChoice trials: We will assess ACP adult host selection and oviposition preference for the same five citrus cultivars, in a pairwise fashion. Potted citrus plants of each cultivar will be pruned to a singled dominant stem, then fertilized to encourage production of flush. A pair of plants of different cultivars will be introduced into a cage, followed by 20CLas-free ACP adults. Plants will be inspected visually daily to carefully count the number of ACP without disturbing them. After 1 week, plants will be inspected to count the number of eggs and nymphs. Adult, egg, and nymph counts on each cultivar will be analyzed with separate generalized linear mixed-effects models (GLMM; Pinheiro & Bates 2000) to assess feeding and oviposition preference among cultivars.Obj. 3aCold chamber study to measure impact of cold on CLas: Young (1-2 year), grafted potted plants of cold-hardy citrus; Tango,Owari, Sugar Belle, Meyer Lemon, Cara Cara, Pink Frost, and Valenciaon two rootstocks (Rubidouxand US-942) will be maintained. We will graft three infected leaves to each receptor and test forCLas at 100 days post inoculation to ensure sufficient infection among the receptor plants. The source will be an infected HLB symptomatic tree, confirmed by qPCR for CLas presence. Uninfected control plants will be included for comparison to monitor impacts ofCLas infection. Once the young plants have developed the disease, plants will be placed in a walk-in freezer room at a range of different temperatures (10, 5, 2, 0 and -2, and -5°C) for 2 h, 4 h, 8 h, 24 h and 72 h duration. CLas titers in citrus will be compared before and after each cold treatment. The experiment will be fully randomized with 4 replicates of each treatment. Analysis will be via a GLM with means separation using Tukey-Kramer test (α = 0.05).Obj. 4aEpigenetics of cold-tolerance in ACP: ACP adults will be placed within a refrigerated climatic chamber to a constant temperature of -4 C° for 10h. Mortality will be assessed after the exposure period. To test for the transmission of the cold tolerance phenotype to offspring through epigenetics, psyllids will be separated into 3 treatments: 1) untreated control, 2) gradual cold acclimation, and 3) intermittent cold acclimation (Martini et al., 2022). Psyllids in the gradual cold acclimation treatment will be isolated in an incubator at 24°C. The temperature will be decreased by 3°C three times a week for 2 weeks until reaching 6°C. In the intermittent cold acclimation treatment, psyllids will be exposed to an overnight drop of temperatures from 21 °C to 6°C, three times a week for 2 weeks. After these cold acclimation treatments, the treatment groups will be put back at 22°C for 3 days and transferred on a small citrus with young flush for oviposition. The nymphs that will emerge from the eggs (generation F1) will be reared to adulthood. Those adults will be tested for cold resistance for 10 h at -4°C, and survivorship will be compared to the control group that will be kept at 22°C for more than 2 generations.Obj. 4bMolecular mechanisms of cold tolerance in ACP: ACP will be attached to a thermocouple coated with a thin film of petroleum jelly and exposed to -1 ° C with a temperature drop of 1 ° C/ h until the latent heat of crystallization that indicates ice formation is detected. Transcriptomic analyses will be conducted on ACP from 1) the ACP lab colony non exposed to any cold acclimation treatment, and 2) ACP after graduate acclimation. For each of these treatments, transcriptomics will be conducted before and after ACP exposure of 10 h at 4 °C. There will be 10 replicates for each treatment, cold exposure, and tissue combination (n=120). Each cDNA library will be sequenced twice using the NovaSeq X Plus Sequencing System. Gene ontology annotation for mapped transcripts will be achieved using Blast2GO and the NCBI database. Individual genes will be considered differentially expressed if the absolute fold change will be ≥ 2 and if the P-value adjusted for false discovery rate < 0.05.Obj. 5a Modeling dynamic risk factors for ACP and HLB spread: The dynamic risk factor model will incorporate natural and human-mediated risk factors across different locations and infrastructure related to citrus production and distribution.These factors facilitate the spread of ACP and HLB through human activities. Spatial analysis will be employed to evaluate risk levels across different regions, providing a comprehensive understanding of spatial risk distribution. Predictive risk mapping will be utilized to provide insights into how changes in environmental conditions or human activities might influence the spread of ACP and HLB.Obj. 5bDeveloping climate suitability model for cold-hardy citrus region: We will gather high-quality climatic data relevant to ACP and HLB distribution from NOAA and Open Meteo API. The data will be preprocessed to standardize formats, eliminate inconsistencies, and ensure compatibility. The model will generate predictive maps highlighting areas of varying suitability for ACP and HLB under current climate conditions. The final model will be integrated with GIS tools to create user-friendly maps and datasets for stakeholders. The model will be accessible through our online tool providing a dynamic resource for future ACP and HLB management in the targeted regions.