Grant Information
| Knowledge Area | Subject of Investigation | Field of Science | Percent |
|---|---|---|---|
| 212 - Pathogens and Nematodes Affecting Plants | 910 - Grapefruit | 1040 - Molecular biology | 50% |
| 212 - Pathogens and Nematodes Affecting Plants | 920 - Orange | 1060 - Biology (whole systems) | 30% |
| 212 - Pathogens and Nematodes Affecting Plants | 2110 - Ornamental trees and shrubs | 1040 - Molecular biology | 20% |
To dissect the disease complex of citrus Huanglongbing (HLB) in Florida from the following five angles:
Through molecular biology techniques, including state-of-the-art genomic sequencing, microarrays, and library construction, we will finish sequencing the Las genome, perform comparative genomics, seed transmission studies and microbial ecology analyses. Using a combination of long established and newly developed microbiology techniques, we will continue to pursue the culturing of Las. Greenhouse and lab experiments will be conducted to monitor Las population dynamics and when appropriate for the progress of other objectives.
To dissect the disease complex of citrus Huanglongbing (HLB) in Florida from the following five angles: 1) to complete Las genome sequence and to conduct comparative genomics of the Liberibacter species; 2) to validate Koch�s postulates by culturing the Las bacterium in vitro and development of artificial inoculation methods; 3) to explore the potential role of the microbial community and genetic diversity of Las bacteria in HLB development; 4) to confirm if Las bacteria are seed-transmissible and their role in HLB development; and 5) to monitor Las population dynamics in citrus and periwinkle plants after chemical and heat treatment. This research relates to inhouse project objective: 1. Characterize ecology, biology, epidemiology, molecular genetics, and vector and host (crop and weed) interactions of domestic, exotic, newly emerging, and re- emerging pathogens, and 2. Develop/refine rapid, sensitive reliable detection/sampling methods for pathogens. Complete genome sequence of Candidatus Liberibacter asiaticus (Las) was obtained using metagenomic approaches with shotgun sequencing and Bacterial Artificial Clone (BAC) libraries. In collaboration with Dr. Hong Lin at the USDA-ARS in Parlier, California, we have also obtained a complete genome sequence of Ca. L. solanacearum. Based on the variations within the Las prophages, FP1 (CP001677.5) and FP2 (JF773396.1), twelve (A to H) different populations (genotypes) have been identified with type A and B localized to FP1 and FP2, respectively. Typing results revealed A, B and C as the three most abundant groups in libraries from psyllid, citrus and periwinkle, although psyllid contained much more type A sequence than either plant host. The various Las populations exist not only in different varieties, but also in different individuals of the same variety. Using Las-specific molecular markers, we are now able to differentiate the Las populations with different pathogenicity, and differentiate psyllid-transmissible and non-transmissble populations. Using PhyloChip� G3, A total of 7,028 known Operational Taxonomic Units (OTUs, "species") of bacteria were detected in citrus leaf midribs of antibiotics-treated and non-treated HLB-affected trees. These OTUs were from 58 phyla with the following five containing 100 or more OTUs: Proteobacteria (44.1%), Firmicutes (23.5%), Actinobacteria (12.4%), Bacteroidetes (6.6%) and Cyanobacteria (3.2%). The results illustrate that the low Las level seen was a combination of a seasonal fluctuation, part of the bacterial population dynamics, and a response to the antibiotic treatments. We have also conducted a few functional genomics studies of the Las bacterium using alternative expression systems to: 1) characterize the Adenosine-5'-triphosphate (ATP) translocase; 2) reveal only one of the two putative znu operons is responsible for zinc uptake; and 3) demonstrate that Las encoded a functional flagellin characteristic of a Pathogen-Associated Molecular Patterns (PAMP).