LIBRO-CONGRESO-CITRUS

lime were treated with chitosan (CHI) 4 mg.mL-1 in HCl 0.05 N pH 5.6. After 24 h, plants were artificially
inoculated with suspension of Xf 9a5c strain (108 cells.mL-1) in PBS buffer or PBS buffer only. The total RNA of
four replicates per treatment (CHI Xf, HCl Xf, CHI PBS and HCl PBS) was extracted and mixed forming a pool of
RNAs which were sequenced in Illumina platform. After sequencing, the transcripts quantitation was used to
calculate the level of differential expression between treatments and control (HCl PBS) and their significance
in Cuffdiff software. We obtained 640 differentially expressed transcripts to CHI treatment, 1232 to CHI Xf and
1075 to HCl Xf. For genes of plant defense we found predominantly repression of genes of auxin pathway. For
the induced genes, CHI treatment showed predominance of genes of flavonoid pathway and hypersensitivity
response, for the CHI Xf were observed predominance of genes of ethylene and jasmonate pathway and for
the HCl Xf genes of salicylic acid pathway, ethylene and jasmonate pathway were observed. Therefore, we
suggest that chitosan induce jasmonate and ethylene-dependent defense-signaling pathway in sweet orange.
Support: INCT Citros (FAPESP and CNPq)
S12P35
Genetic variation of California Spiroplasma citri populations revealed by two genetic loci
Wang X. 1 , Doddapaneni H. 2 , Chen J.C. 1 , and Yokomi R.K. 1
1 USDA Agricultural Research Service, San Joaquin Valley Agricutural Sciences Center (USDA ARS SJVASC), Crop Diseases, Pests and
Genetics, USA; and 2 Baylor College of Medicine (BCM), Human Genome Sequencing Center, USA. ray.yokomi@ars.usda.gov
Citrus Stubborn Disease (CSD), known to be present in California since 1915, was confirmed to be caused
by Spiroplasma citri in 1972. Hosts of S. citri include citrus and a wide range of annual weeds, ornamentals
and crops such as carrots and sesame. Genetic variation of S. citri in California was examined previously by
RAPD-PCR with 20 primer pairs but no unique genetic signatures were found. Using partial chromosome
and plasmid sequences of S. citri recently released in GenBank, a conserved hypothetic peptidyl-arginine
deiminase protein (PADP) and a transmembrane protein (TMP) of plectrovirus spv1-r8a2b were selected in
silico to evaluate genetic diversity of S. citri from different hosts and locations. Two specific PCR primer sets
designed proved to be effective. The first was PADPf/PADPr which targeted a region with variable tandem
repeat numbers (TRNs); the second was TMPf/TMPr which had multiple priming sites in the published
genome of S. citri strain GII3-3X. A panel of 31 strains of S. citri from California, Illinois and the Mediterranean
region was evaluated using the two primer sets by PCR. Electrophoretic profiles of PCR amplicons from
the strains tested consistently showed three TRN patterns in the PADP locus and three in the TMP locus.
Interestingly, strains which showed a different profile for PADP also showed a different profile for TMP and
each pattern group constituted a set. Sequence analyses were conducted amongst strains in each pattern
group and revealed at least three genotypes present in Californian S. citri populations. Further study of
field strains from different geographical origins using the two loci described should provide insight on the
biology and epidemiology of S. citri.
S12P36
Serological detection of Spiroplasma citri using a bacterial secreted protein as the detection
marker
Shi J.S., Pagliaccia P.D., Morgan M.R., Ma W.M., and Vidalakis G.
University of California, Riverside (UCR), Plant Pathology and Microbiology, U.S.A. vidalg@ucr.edu
Citrus worldwide is facing major threats from insect-transmitted and phloem-limited bacterial diseases such
as Huanglongbing (HLB, “Candidatus Liberibacter” spp.) and Stubborn (CSD, Spiroplasma citri). Management
of such diseases is difficult, expensive, heavily based on tree eradication and insecticide treatments for vector
control. The success of any disease management program is based on early pathogen detection. The most
commonly used diagnostic or survey methods are PCR based and require the presence of bacterial cells or
DNA in the tested sample for positive diagnosis. This can be problematic in the case of citrus since only a few
leaves or stems from a whole tree and eventually a few mg of tissue are processed for DNA extraction while
the titer and distribution of the pathogen within trees, types of tissue and disease progression are variable.
XII INTERNATIONAL CITRUS CONGRESS 2012 - 213
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