Progress 08/25/05 to 08/01/07
Outputs
Progress Report Objectives (from AD-416) The objectives of this cooperative research project are to develop a database of genomics information related to citrus pathogens (citrus tristeza virus and Xylella fastidiosa), infection and citrus pathogen- citrus host interactions for use in developing rapid, reliable microarray- based citrus pathogen or pathogen infection detection systems and novel therapeutic approaches for citrus disease management. Approach (from AD-416) A genomics database including (but not necessarily limited to) genome sequences, annotations and ESTs related to Xylella fastidiosa (Xf) and citrus tristeza virus (CTV), as well as Xf-/CTV-host interactions will be developed based on available information from various sources. Bioinformatic tools will be used to mine this information to identify and select Xf- and CTV-specific genomic signatures, as well as EST sequences responsive to Xf and CTV infection. Information gaps to develop the microarray-based detection systems and novel therapeutic tools will be identified. The database and bioinformatics information will be disseminated to cooperating state and federal scientists, and to interested stakeholders. Documents Non-funded Cooperative Agreement with Citrus Research Board. Formerly 5302-22000-007-16N (3/07). Significant Activities that Support Special Target Populations This report documents research under a non-funded Agreement between ARS and the California Citrus Research Board. Additional details of the research can be found in the report for the in-house associated project 5302-22000-008-00D Epidemiology and Management of Xylella fastidiosa (Xf) and Other Exotic and Invasive Diseases and Insect Pests. The research relates to the in-house project objective to develop effective, environmentally-sound methods to manage, and mitigate the effects of citrus diseases, including citrus canker (CBCD), citrus variegated chlorosis (CVC), citrus tristeza and huanglongbing (HLB). The overall goal was to develop a comprehensive bioinformatics data warehouse of citrus pathogens and pests to aid in the disease early diagnosis and pathogen detection. The specific objective was to maintain and update the genomic information as well as conduct comparative analyses for the development of microarray-based pathogen detection systems. The citrus pathogens targeted for this project were Xylella fastidiosa, Xanthomonas axonopodis pv. citri, citrus tristeza virus, stubborn and Candidatus Liberibacter species, as well as plant parasitic nematodes and insect pests. Whole genomic sequences and ribosomal RNA sequences of 16S, 23S, 18S, 5.8S and their spacer sequences were used to construct the database. Sequence alignment files are provided when possible. The Citrus Disease Database (CDD) has a dedicated domain name (http://cropdisease.ars.usda. gov) and IP address: (http:// 199.133.142.23). This database is currently located on an ARS server in Parlier, CA, includes available genomics and bioinformatics information of 36 species or 42.6% of total citrus pathogens (bacteria, fungi, viruses), plant parasitic nematodes, and insect pests. Data files containing genomic sequences of citrus pathogens, plant parasitic nematodes and insect pests are publicly accessible and downloadable from the CDD database. More than 1,700 published articles regarding the citrus pathogens (HLB, CVC, CTV, Xac, Stubborn, insects, plant parasitic nematodes) are incorporated into this database. Two bioinformatics tools (SCPrimer and nWayComp) were developed and are downloadable from the CDD. A branch, grape EST database called VitisDB (http://cropdisease.ars.usda.gov/~fruit_tree) was also developed. It contains grape (V. rupestris � V. arizonica) expression data related to X. fastidiosa. This database will contribute to (1) development of improved, rapid, reliable microarray-based citrus pathogen or pathogen infection detection systems and (2) novel therapeutic approaches for citrus disease management. Activities and progress related to this cooperative project were monitored by review and discussion of results through regular conversations daily or weekly and co-preparation of presentations of research reports at annual stakeholder and scientific meetings.
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Progress 10/01/05 to 09/30/06
Outputs
Progress Report 4d Progress report. This report serves to document research conducted under a Specific Cooperative Agreement between ARS and the Citrus Research Board, Visalia, California. Additional details of the research can be found in the report for the parent project 5302-22000-007-00D, Epidemiology and Management of Xylella fastidiosa (Xf)-caused Diseases and Xf Insect Vectors. Databases of four citrus pathogens (Xylella fastidiosa, Xanthomonas spp., Citrus tristeza virus and Candidatus Liberibacter species) were established. They are named CVC-database [1], Canker-database [2], CTV-database [3] and HLB-database [4]. CVC-database was based on sequences of four available Xf genomes. Analysis of the genome-wide variations among multiple Xf strains has been demonstrated. The Canker-database consists of the available sequences of the genome sequences of one Xac (X. axonopodis pv. citri str. 306) and five other plant pathogenic xanthomonad strains. The comparative
genome sequences of the six xanthomonad strains have been completed. The utility of the CVC-database and Canker-database was demonstrated by designing specific primers for Xf- CVC and Xac for the development of new or novel diagnostic platforms (e.g. , multiplex citrus pathogen DNA microarray chip). CTV-database contains whole genome sequences of eight strains. In addition, the partial genome sequence of 211 isolates (strains), 853 entries of nine major genes of CTV were added. Also included are detection primers and 13 entries of 5 UTR sequences. Limited genomic sequence information of Ca. Liberibacter species is available. A total of 64 genomic sequences, regarding the 16S rDNA sequences, outer membrane protein and sequences encoding ribosomal genes of the beta-operon, have been collected in HLB-database. The sequence information can be retrieved in the order of their Genbank GI number, strain type, length, date of submission, collection loci, submission date, function description
and submission institute. Adatabase of grape and citrus expression GO ontology database [5] (EST and microarray) has been developed. Primarily it contains the grape expression EST GO terms after inoculation with Xylella fastidiosa and citrus expression EST GO terms after inoculation with multiple pathogens. In addition to the above database construction, two bioinformatics tools were developed: SCPrimer [6] and nWayComp [7]. SCPrimer is used to design specific primer for pathogen detection at genome scale level. nWayComp is used for the sequence comparison and therefore identify unique genes (sequences) for pathogen detection. The citrus pathogen database has been made available to the National Plant Pathogen Consortium through the Los Alamos National Laboratory in Los Alamos, New Mexico. The database is updated continuously. The information can be found at: 1.CVC-database: http://fresno.ars.usda.gov/citrus-disease/CVC_index.htm; 2.Canker-database:
http://fresno.ars.usda.gov/citrus-disease/Canker_index. htm; 3.CTV-database: http://fresno.ars.usda.gov/citrus- disease/Canker_index.htm; 4.HLB-database: http://fresno.ars.usda. gov/citrus-disease/greening_index.htm; 5.GO-database: http://cropdisease.ars.usda.gov/~fruit_tree; 6.SCPrimer: http://fresno.ars.usda.gov/citrusdisease/downloads/; 7.nWayComp: http://fresno.ars.usda.gov/citrusdisease/downloads/nwaycomp.htm
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