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United States Department of Agriculture

Agricultural Research Service

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Research Project: Mini-Plant Indexing: Development of Protocols to use Very Young Plants for Biological Indexing

Location: National Clonal Germplasm Repository for Citrus

2013 Annual Report

1a. Objectives (from AD-416):
This is a project to further examine the use of young plants for biological indexing: to develop a standardized protocol, determine the best potting medium (UC Mix may not be the optimal for conetainers), determine the method of grafting, explore the possibility of placing the indictor plants, once inoculated, in an environmental chamber under optimum temperature to get quicker symptom development. Development of standardized protocols and the ability to conduct biological indexing with plants that can be grown and used in weeks instead of months and in significantly less space would enable the California Nursery Industry to continue recurring biological indexing for psorosis, tristeza, and viroids for mother trees, thus providing better protection for the industry as compared to increased reliance on laboratory testing where only target pathogens would be found, but not unknown pathogens.

1b. Approach (from AD-416):
The overall objective is to evaluate the potential of using very young plants in conetainers or small pots for performance in biological indexing for the graft transmissible pathogens of citrus. This will be done by comparing the results of biological indexing using young plants, traditional biological indexing approach and the use of laboratory methods. A standardized protocol will be developed after considering different size conetainers and potting systems, comparison of potting medium, evaluate of methods of inoculation of small plants for consistency of performance and inoculum survival, evaluation of the potential of using environmental chambers or Arabidopsis chambers, examining economical methods to maintain desired temperatures. Once the protocol has been standardized, a workshop will be held for other certification programs, nurserymen, and other interested parties to demonstrate the methodology.

3. Progress Report:
This project is related to objectives 1 and 3 (identification of priority genetic resources of citrus and develop more accurate and high through-put diagnostic methods for priority graft transmissible pathogens of citrus, respectively). Citrus germplasm must be pathogen-tested for and found free of all known graft transmissible pathogens before being released from quarantine status and available for distribution and registered budwood source trees must be re-indexed at regular intervals. The California Department of Food and Agriculture requires most of the pathogen-testing to be performed by biological indexing using indicator plants, selected for use because they produce disease symptoms when infected with specific graft transmissible pathogens and incubated at the correct temperature for development of disease symptoms. A standard protocol has been developed for biological testing of citrus pathogens, beginning with the inoculation of 8-12 month old indicator plants. In this project, we are evaluating the performance of very young indicator plants, grown for 60-75 days from seed, and planted in small containers to minimize bench space. We have demonstrated the ability to detect graft transmissible pathogens requiring cool temperature conditions, including Citrus psorosis virus, concave gum disease, Dweet mottle virus, and Citrus tatterleaf virus, during the summer months by using very young indicator plants placed near the evaporative cooling pads in the greenhouse. Results obtained were the same when compared to biological indexing of the same source plants indexed during the winter months using the standard protocol. Application of mini-plant indexing for use in citrus nurseries in California will reduce the need for quality greenhouse space required for growing standard indicator plants for 8-12 months, enable biological indexing to be performed year round at a reduced cost, and help ensure citrus nurseries have only healthy plants.

4. Accomplishments

Last Modified: 06/23/2017
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