Host pathogen interaction, and copper resistance in Xanthomonas associated with citrus canker

Abstract

Citrus canker is a bacterial disease caused by Xanthomonas citri type A (Xc-A) and X. fuscans pv. aurantifolii (Xfa). The disease reduces yield and affects marketing of the fruits. Most of the strategies for citrus canker control are based on application of copper sprays. Two strains, Xc-Aw and Xfa type C (Xfa-C), isolated in Florida and Brazil, elicit a hypersensitive reaction (HR) in grapefruit (Citrus paradisi) leaves. The major objectives of this dissertation are: 1) to identify and characterize the gene from Xfa type C that elicits HR in citrus species, and 2) to characterize chromosomal and plasmid rearrangements in copper resistant (CuR) strain Xc-A44. The gene responsible for the Xfa type C strain eliciting an HR in citrus species except Key lime (C. aurantifolia) was identified and designated avrGf2. A non-functional copy of this effector is present in Xfa-B strains. Compared with the previously characterized gene avrGf1, avrGf2 elicited a faster form of HR in citrus. The two avirulence genes, avrGf1 and avrGf2, both members of the XopAG family, have 45% identity at the amino acid level and contain chloroplast localization signals. Based on amino acid sequence comparisons, two C-terminal motifs were identified in AvrGf1 and AvrGf2 effectors and also in several XopAG already described. The effector AvrGf2 shows a putative C-terminal domain (CLNAxYD), and a cyclophilin-binding site (GPxL), which was determined to be essential for HR elicitation. The genome sequence of strain Xc-A44, shows a similar T3E profile as well high sequence identity with the chromosome of the reference strain Xc-A306, although there are considerable differences in their plasmids. The first plasmid difference is associated with the pathogenicity plasmid, which has an approx. size of 123 Kbp. Plasmid Xc-A44p123 has differences in the sequence and number of its TALEs; this plasmid contained two functional copies of pthA4 gene, and nucleotide sequences that resemble a recombination of plasmids pXAC33 and pXAC64 from strain Xc-A306. The second plasmid (pCuR) has two sequence insertions with homology to the Xc-A44 and Xc-A306 chromosome. The insertions found in Xc-A44pCuR have sequence identities with transposon genes previously identified in several Xanthomonas species.