Location: National Clonal Germplasm Repository for Citrus
2020 Annual Report
Accomplishments
1. Support of critical research on Huanglongbing. Support of critical research on Huanglongbing. Huanglongbing (HLB) remains a threat to commercial citrus production. Researchers at ARS, land grant universities, and other entities are actively working on solutions. The ARS Citrus and Date Repository in Riverside, California, supplies citrus germplasm to researchers in Ithaca, New York; Wimauma, Fort Pierce, and Lake Alfred, Florida; New Haven, Connecticut; West Lafayette, Indiana; Beltsville, Maryland; and elsewhere supporting HLB-related research. This germplasm is both tested for susceptibility and used for making crosses to create new Citrus hybrids with greater resistance to HLB.
2. Provides clean source citrus propagative material. As a component of the National Clean Plant Network – Citrus, ARS researchers in Riverside, California, supplied clean propagative materials to the minor citrus producing states that do not have a State-level program (Arizona, Louisiana, Alabama, Georgia). ARS in Riverside, California, also provided clean source propagative materials to programs in other states and countries. This material is used for making crosses to develop improved cultivars. This will improve the overall plant health of the global citrus industry.
3. Evaluation of early detection technologies for huanglongbing (HLB)-associated pathogens. Evaluation of early detection technologies for huanglongbing (HLB)-associated pathogens. If HLB can be detected early, infected trees can be removed quickly, which will slow the spread of this disease and help save the Citrus industry. Early detection technologies (EDT) investigated include: HLB detection canines, serological assays for proteins secreted by the pathogen and bacterial structural proteins, changes in the metabolomic profile of the host plant, and next generation sequencing (NGS). These methods were compared to the standard regulatory quantitative Polymerase Chain Reaction (qPCR) diagnostic method. While the EDT’s have identified suspect positive trees, extensive qPCR assays have negative results. Canines continue to be the most rapid detection method and research continues, to determine the time between canine detection and a positive qPCR result for the same tree.
4. Development of tissue blot method for citrus pathogen detection. Development of tissue blot method for citrus pathogen detection. Citrus germplasm collections in both the field and within protective structures are routinely screened for pathogens. Nucleic acid extraction kits are costly. A forty-nine percent reduction in cost of supplies was realized by blotting fresh bark tissues onto nitrocellulose membranes, cutting out the blots, and eluting the nucleic acids from the membrane. Quantitative polymerase chain reaction was performed using the crude nucleic acid extraction. This method has proven accurate for the detection of citrus viroids and huanglongbing, which is necessary to determine if the collection remains free from these pathogens.
5. Selection of pigmented Australian finger limes. Selection of pigmented Australian finger limes. Pigmented seedlings of Australian finger limes were evaluated by scientists in Riverside, California, for horticultural characteristics in anticipation of therapy and release from quarantine in the next few years. Currently, only one lightly pigmented selection is available to stakeholders and the industry desires more options.
6. Estimation of actual water use in date palm production. Estimation of actual water use in date palm production. In cooperation with scientists in Riverside and Imperial, California, actual water consumption of mature date palms was measured using various techniques. Results indicate that although there is some variability in date palm water use, actual use is substantially lower than water applications by many growers. As water supplies become more restricted in the desert area, this information will be valuable in conserving water in date production.
7. Development of an array for simultaneous detection of 15 RNA viruses and viroids. Development of an array for simultaneous detection of 15 RNA viruses and viroids. A novel array method was developed for the simultaneous detection of two targets each from 15 different citrus ribonucleic acid (RNA) pathogens in addition to two reference gene targets. The 96 well assay plate can be used for assaying two plant samples (RNA extractions) along with one pooled positive control RNA sample in about 75 minutes. The array consists of 32 sets of primers and probes along with stabilizers and reporters placed in individual wells, lyophilized and stored frozen. The platform was used to detect actual pathogen status (15 RNA pathogens) of about 50 accessions in the citrus pathogen inventory. This method was validated by the ARS, Riverside, laboratory personnel in independent tests.
8. Use of Citrus tristeza virus (CTV) as a vector for antimicrobial peptides and ribonucleic acid interference (RNAi). Use of Citrus tristeza virus (CTV) as a vector for antimicrobial peptides and ribonucleic acid interference (RNAi). Similar to the Citrus tristeza virus (CTV)-vector developed in Florida, mild California CTV isolates collected by the Central California Tristeza Eradication Agency were cloned and transformed by collaborating scientists at the University of California - Riverside. ARS scientists and technicians provided healthy citrus plants, inoculated plants with purified virus, and assayed for CTV. After three years, a stable CTV vector was developed and is now ready to use as a delivery mechanism for therapeutic antimicrobial peptides and RNAi strategies against pathogens such as Huanglongbing-associated bacteria and insect pests.
Review Publications
Hazzouri, K.M., Gros-Balthazard, M., Flowers, J.M., Copetti, D., Lemansour, A., Lebrun, M., Masmoudi, K., Ferrand, S., Dhar, M.I., Fresquez, S., Roas, U., Zhang, J., Talag, J., Lee, S., Kudrna, D., Powell, R.F., Leitch, I.J., Krueger, R., Wing, R., Amiri, K.M., Purugganan, M.D. 2019. Genome-wide association mapping of date palm fruit traits. Nature Communications. https://doi.org/10.1038/s41467-019-12604-9.
Ollitrault, P., Curk, F., Krueger, R. 2020. Citrus taxonomy. Book Chapter. https://doi.org/10.1016/B978-0-12-812163-4.00004-8.
Ramsey, J.S., Chin, E., Chavez, J., Saha, S., Mischuk, D., Mahoney, J., Mohr, J., Robison, F., Godfrey, K., Levesque, C., Foster, L., Xu, Y., Strickler, S., Fernandez, N., Polek, M., Giovannoni, J.J., Mueller, L., Slupsky, C., Bruce, J., Heck, M.L. 2020. Longitudinal transcriptomic, proteomic, and metabolomic analysis of Citrus limon response to graft inoculation by ‘Candidatus Liberibacter asiaticus’. Journal of Proteome Research. 19:2247-2263. https://doi.org/10.1021/acs.jproteome.9b00802.
Salomon-Torres, R., Sol-Uribe, J., Valdez-Salas, B., Garcia-Gonzalez, C., Krueger, R., Hernandez-Balbuena, D., Norzagaray-Plascenci, S., Garcia-Vazquez, J., Ortiz-Uribe, N. 2020. Effect of four pollinating sources on nutritional properties if ‘Medjool’ date (Phoenix dactylifera L.) seeds. Agriculture 2020, 10 (45). https://doi.org/10.3390/agriculture10020045.
Gottwald, T.R., Luo, W., Posny, D., Poole, G.H., Louws, F., Mccollum, T.G., Hartung, J.S., Bai, J., Duan, Y., Taylor, E.L., Da Graça, J., Schneider, W., Polek, M., Hall, D. 2020. Canine olfactory detection of a vectored phytobacterial pathogen, Liberibacter asiaticus, and intergration with disease control. Proceedings of the National Academy of Sciences. 117(7)3492-3501. https://doi.org/10.1073/pnas.1914296117.
Padhi, E.M., Maharaj, N., Lin, S., Mishchuk, D.O., Chin, E., Godfrey, K., Foster, E., Polek, M., Leveau, J.H., Slupsky, C. 2019. Metabolome and microbiome signatures in the roots of citrus affected by Huanglongbing. Phytopathology, 109 (12). https://doi.org/10.1094/PHYTO-03-19-0103-R.
