1a. Objectives (from AD-416):
To produce citrus scion genotypes which are resistant to huanglongbing (HLB) using conventional plant breeding and on-transgenic biotechnology.
1b. Approach (from AD-416):
In the USDA breeding program several hybrids display tolerance to huanglongbing (HLB), with plants testing positive for Liberibacter but continuing to grow and crop well. Among commercial cultivars, Temple has the best HLB-resistance in initial tests. In this study, we will quantify the HLB-resistance/tolerance of priority material to determine its value in commercial Florida citrus production. We will also develop tissue culture derived material in which resistant genotypes form the vascular system and standard varieties produce the fruit quality layers. Such chimeras exist as commercial cultivars in China. Like scions on rootstocks, these chimeras use genotypes for their best traits in each plant portion, with proven methodology. The tissue culture "internal grafting" procedure to produce periclinal chimeras was developed by Ohtsu (Ann. Phytopath. Soc. Japan 60:82-88) and further demonstrated by Sugawara et al. (HortScience 30:1276 -1278) in Japan. Since the tissue layer forming the phloem and xylem contribute little to the juice vesicles, use of an HLB-resistant internal layer may provide a rapid and sustainable solution to HLB using established fruiting types. Material displaying promise will be further tested for fruit quality and other characteristics, while being field tested against standards to assess HLB resistance.
3. Progress Report:
This project is related to Objective 1: Create new genetic combinations of citrus, Objective 2: Screen germplasm for important traits and select superior individuals, Sub objective 1 D: Create new scions and rootstocks with potential resistance to huanglongbing (HLB) and citrus bacterial canker (CBC) by genetic transformation. Evaluation of existing standard cultivars (‘Temple’, ‘Fallglo’, ‘Sugar Belle’, ‘Tango’, ‘Hamlin’, and ‘Ruby’) for huanglongbing (HLB) tolerance/resistance is underway. Trees were planted in 2010, using a randomized complete block design, at Picos Farm, Ft. Pierce, FL. huanglongbing symptom development and tree growth (diameter and height) are being monitored on a monthly basis. All of the cultivars in this trial exhibit symptoms of huanglongbing and have tested positive for Candidatus Liberibacter asiaticus Results to date support earlier observations that ‘Temple’ and 'Fallglo' are in the most tolerant group. Anatomical features associated with tolerant vs. susceptible closely related genotypes are being investigated. Electron micrographs and visual images are being compared. Fourier transform infrared spectroscopy, which can be focused on specific tissues, displays promising differences between resistant and susceptible material. Another project involves the treatment of various resistant/tolerant citrus accessions and susceptible standards with various concentrations of antibiotics to generate a range of Candidatus Liberibacter asiaticus titer levels. This has been slightly delayed and in May 2013, budwood with various concentrations of Candidatus Liberibacter asiaticus, derived from the antibiotic treated plants, will be evaluated for their potential to result in huanglongbing symptoms in disease free material. The budded plants will be evaluated for growth and huanglongbing symptoms development over a 2-year period. Temporal progression and systemic movement of the bacteria in the inoculated plants will be determined along with huanglongbing symptom development, and growth of the plants. Development of periclinal chimera using resistant genotypes and standard varieties is in progress. In vitro shoots have been established from nodal and internodal explants excised from mature, certified disease free plants of Red Carrizo, Temple, Hamlin, and Valencia. After root formation, chimeras will be generated. After successfully generating the chimeras with huanglongbing resistant vascular system and good fruit using the previously mentioned cultivars, additional cultivars such as ‘Sweet Orange’ and grapefruit will be added to this study. One additional study has been added to the project. Screening and evaluating new scion materials is a lengthy process and require multiple testing locations. Due to the urgency to develop tolerant/resistant material, a shorter evaluation cycle procedure is being investigated. If this screening method is successfully, it may be useful to quickly identify new sources of resistance varieties that may enhance and improve citrus production in Florida.