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ARS Home » Southeast Area » Fort Pierce, Florida » U.S. Horticultural Research Laboratory » Citrus and Other Subtropical Products Research » Research » Research Project #437845

Research Project: Sustainable Citrus Production

Location: Citrus and Other Subtropical Products Research

2020 Annual Report

Objective 1: Determine genetic and horticultural inputs that provide superior citrus production and quality under endemic HLB disease pressure. [NP 305, Component 1,Problem Statement lB] • Sub objective 1a. Evaluate susceptibility of standard and novel citrus scion and rootstock combinations in grower-cooperator trials. • Sub objective 1b. Determine the effects of prophylactic treatments on prevention of HLB. • Sub objective 1c. Determine effects of pre-plant soil treatments on citrus tree health and productivity. Objective 2: Investigate plant physiology and symptoms during disease development to design rational approaches for treatment. [NP 305, Component 1, Problem Statement lB Sub objective 2a: Determine the effects of CLas infection on photosynthesis and carbon partitioning in citrus.

This project includes field plot, greenhouse and laboratory components to evaluate factors related to citrus susceptibility to Huanglongbing disease. Field plots will be conducted with grower cooperators to evaluate performance of scion/rootstock combinations for susceptibility to Huanglongbing disease under commercial management systems. Some such trial were established in the previous project and data collection will continue in the current project. New plantings will also be established in the current project. Citrus is a long term crop that does not begin to produce fruit until three to four years after planting. During the first three years of the experiments, tree mortality is really the only metric for tree performance. When trees come into bearing yield and fruit quality data will be collected. Greenhouse trials will be conducted to determine the efficacy of prophylactic treatments to prevent Haunglongbing disease. In the previous project realistic model system including citrus, Asian citrus psyllids and Candidatus Liberibacter asiaticus. This model has a small foot print and is rapid, and robust. Numerous therapeutic treatments to mitigate Huanglongbing symptoms, but there have been none of these treatments have proven efficacious. In this project trees will be treated prior to transmission of Candidatus Liberibacter asiaticus by Asian citrus psyllids. The hypothesis is that preventing the establishment of Candidatus Liberibacter asiaticus will be effective whereas therapeutic treatments have unsuccessful. Data will be collected to determine the effects of treatments on incidence of Candidatus Liberibacter infection incidence and titer, growth and development of Huanglongbing symptoms. If promising treatments are identified in the greenhouse, small scale field trails will be established to determine efficacy of these treatments in the field. Grower cooperator trials will also be established to determine if anaerobic soil disinfestation, a treatment that alters the soil microbiome and has proven beneficial for some annual crops and may improve the growth of newly planted citrus trees. Data collection will be as described for the scion/rootstock evaluations. Greenhouse trials will also be conducted to determine how citrus leaf ontogeny relates to the development of Huanglongbing symptoms. It is apparent that it is only the very youngest of citrus leaves is where transmission of Candidatus Liberibacter asiaticus. Anatomical changes that develop very early following Candidatus Liberibacter asiaticus will be investigated using light and electron microscopic evaluation. In addition, because Huanglongbing is characterized by massive accumulation of starch, the effects of Candidatus Liberibacter on photosynthetic activity will be determined by quantifying starch accumulation. The effect of infection on the rate of carbon dioxide fixation (photosynthesis) will be determined using commercially available instrumentation. Carbon partitioning (dry weight) following infection will also be investigated by determining root and shoot dry weights post infection.

Progress Report
Subobjective 1.a. Data was collected on tree health and productivity in most trials, data was collected on fruit quality in trails that produced sufficient fruit. Anaerobic Soil Disinfestation (ASD) trial that was initiated six years previously was completed. Data was collected on tree mortality in a cooperative trial with the University of Florida and a commercial lemon grower in Saint Lucie County. As part of a Multi-Agency Coordination (MAC) Group-funded project worked with University of California, Riverside, scientist to develop plans for testing USDA advanced citrus scion hybrids in field trials in California. This work was contingent on an Material Transfer Agreement (MTA) that allowed for the importation of citrus from Florida into California which was accomplished in the first phase of the project. Subobjective 1.b. Significant progress was made towards this milestone. Five experiments to determine the effects of prophylactic treatments on prevention of Huanglongbing (HLB) disease were conducted and data has been collected and analyzed. Two progress reports on this work were submitted to the Citrus Research Board that provided partial funding for the project. Subobjective 2.a. An ARS technician was sent to LiCor Headquarters for training on the use of the portable CO2 fixation instrument. Initial experiments with the LiCor unit were conducted with citrus in Ft. Pierce, Florida. Operating parameters for the measurement of carbon dioxide fixation and chlorophyll fluorescence. Subsequently, experiments were conducted to determine effects of light level and time of day on carbon dioxide fixation rates. More progress could have been made toward this milestone had it not been for the pandemic. Subobjective 2.b. The time course experiment was conducted, some data and samples for detection of Candidatus Liberibacter asiaticus (CLas) (causal agent of HLB) were collected. Unfortunately, the data collection phase of this experiment was during the pandemic, which prevented the collection of samples for microscopy and carbohydrate analyses. This experiment will have to be repeated. Plant material that was being grown to conduct the experiments was lost due to the pandemic. Additional replications of the experiment need to be conducted.

1. Field plot experiments. Extensive field plot (USDA and commercial cooperator properties) experiments continue from previous years and new plantings were established. Data is being collected to evaluate tree health, with a focus on susceptibility to HLB, in novel citrus scion and rootstock combinations. In trails with bearing trees data is collected on yield and fruit quality. An orchard trial was established with a commercial lemon grower in Saint Lucie County, Florida to determine the impacts of anaerobic soil disinfestation on establishment and early growth of lemons. As part of a MAC-funded project, ARS researchers in Fort Pierce, Florida, in collaboration with University of Florida and University of California established a large (ca. 1,000 tree field trial) to conduct genotypic evaluation of HLB susceptibility. Worked in collaboration with rootstock breeder to determine effects of rootstocks on fruit quality, ca. 2,000 juice samples that were processed. In collaboration with rootstock breeder released “mini finger lime”. As part of a National Institute of Food and Agricultural Science (NIFA)-funded project ARS researchers in Fort Pierce, Florida, collaborated with University of California scientist to evaluate HLB susceptibility of numerous novel citrus hybrids.

2. Greenhouse model for studying the Citrus-ACP-CLas pathosystem. Lack of a model system for the study of HLB has been recognized as a research impediment by citrus scientists and the citrus industry. In previous years of this project a greenhouse model for studying the Citrus-ACP-CLas pathosystem was developed. In the current year, the model system was used in a number of experiments both for the in-house project as well as for several grant-funded projects. Based on results of these experiments new insights have been gained regarding the generational transmission of CLas in ACP, the relationship between CLas titer in ACP and subsequent transmission and HLB development, timing between transmission and appearance of HLB symptoms. All of these variables have fundamental biological significance to HLB epidemics and provide important information for modeling HLB epidemics. Citrus trees produced with the model system have been utilized in numerous “early detection” projects, including one that was published in PNAS. Collaborations between ARS scientists at Ft. Pierce, Florida, and USDA APHIS regarding early detection continue and two experiments were conducted. In a Citrus Research Board-funded project the model system was used to test the hypothesis that antibiotic treatments applied prophylactically to citrus could impact subsequent HLB following exposure to CLas-infected ACP. ARS was approached by the University of California Riverside BLS3 citrus research facility to provide both technical advice as well as CLas-infected citrus for use in their research efforts. ARS has also been approached by agrichemical and agritechnical companies with interest in the model system.

Review Publications
Zuniga, C., Peacock, B., Liang, B., Mccollum, T.G., Irogoyen, S.C., Tec, D., Marotz, C., Weng, N., Zepeda, A., Vidalakis, G. 2020. An in vitro pipeline for screening and selection of citrus- associated microbiota with potential anti-“Candidatus Liberibacter asiaticus” properties. Applied and Environmental Microbiology.
Hunter, W.B., Clarke, S., Sandoval Mojica, A., Paris, T., Miles, G., Metz, J., Holland, C., McCollum, T.G., Qureshi, J., Tomich, J., Boyle, M., Cano, L., Altman, S., Pelz-Stelinski, K. 2020. Advances in RNA suppression of the Asian citrus psyllid vector and bacteria (Huanglongbing pathosystem). Chpt. 17. 352 pages. In Asian Citrus Psyllid. Biology, Ecology and Management of the Huanglongbing Vector. (eds) Phil Stansly and Jawwad Qureshi. Commonwealth Agricultural Bureau International (CABI Press). p. 258-283.
Hong, J.C., Di Gioia, F., Jones, J.B., Turechek, W., Johns, C.W., Finley, N.L., Ozores-Hampton, M., Mccollum, T.G., Rosskopf, E.N., Burelle, N.K. 2020. Defining anaerobic soil disinfestation through changes in the microbiome. Acta Horticulturae.
Ginnan, N.A., Dang, T., Bodaghi, S., Ruegger, P.M., Mccollum, T.G., England, G., Vidilakis, G., Borneman, J., Rolshausen, P.E., Roper, C.M. 2020. Disease-induced microbial shifts in citrus indicate microbiome-derived responses to Huanglongbing across the disease severity spectrum. The ISME Journal: Multidisciplinary Journal of Microbial Ecology. online.