Project description:Xanthomonas campestris pathovar campestris (Xcc), the causal agent of black rot disease of cruciferous plants worldwide, is composed of phenotypically heterogeneous groups of strains. The knowledge about the genome diversity and phylogenetic relationships between Xcc strains with different origins are of great interest as they provide insight into the mechanisms of pathogenicity, host preferences and evolution of this pathogen. In our present work, eighteen Xcc strains collected from different geographical area of China mainland were investigated concerning of the genome composition by comparative genomic hybridization (CGH) using microarray slides spotted with PCR-based intragenic DNA fragments of 4273 open reading frames (ORFs) representing the non-redundant genome content of Xcc strain 8004. The common genome backbone of Chinese strains was estimated to contain about 3404 ORFs, which was considered to maintain the basic characteristics of Xcc, i.e. the yellow mucoid colony on nutrient solid medium as well as the pathogenicity to induce black rot disease on host plants. A flexible gene pool of 729 ORFs in Xcc was characterized, of which 402 ORFs were clustered in twenty-seven highly variable genomic regions in Xcc 8004. Of these highly variable genomic regions, five are absolutely absent from Chinese strains, which constitutes the main genomic differences between the Xcc 8004 and Chinese strains. Transcriptome analysis of Xcc 8004 grown in the rich medium NYG and the defined medium XVM2 indicated that the expression of some certain genes in highly variable genomic regions are significantly activated in XVM2, which included the predicted pathogencity and avirulence genes. Candidate genes for cultivar-specificity of Xcc were identified in the variable genomic regions: the avrXccC and avrXccE1 were demonstrated to confer the avirulence on the host plants Mustard cultivar (cv.) Guangtou and Chinese cabbage cv. Zhongbai 83, respectively; and the avrBs1 showed to correlate with the hypersensitive reaction (HR) on the non-host plant pepper ECW10R. This study revealed the common genome backbone of Xcc maintained the basic function in essential metabolisms and basic pathogencity, and the variable genomic determinants contributed to the cultivar-specificity of the pathogen, suggesting that the Xcc genome, with a compact function core carrying essential genes for survival, reproduction, and invasion, is constantly diversifying by acquiring and losing DNA segments, or by DNA degeneration, to improve the genetic novelty for the adaptation during the evolution. Keywords: CGH analysis and transcriptome analysis

Project description:Xanthomonas is one important model microbe to study the molecular determinants of virulence and host range of pathogens since Xanthomonas is capable of infecting numerous monocotyledonous and dicotyledonous plants. Among the plant diseases caused by Xanthomonas, X. citri subsp. citri (Xcc) causes citrus canker, which has significant impact on citrus production. Xcc is classified into different strains primarily by host range including A and Aw. The A (Asiatic) strain (XccA) has a wide host range and is most virulent, whereas Aw (Wellington) strain has restricted host range including Key or Mexican lime and alemow. We hypothesized that not only gene content but also gene expression contributes to the difference in virulence and host range of closely related strains. To test our hypothesis, comparative genomic and transcriptome analyses were conducted to study the two closely related Xcc A and Aw strains. The genome of X. citri subsp. citri strain Aw12879 (Xcaw) was completely sequenced using 454 Pyrosequencing, Illumina sequencing and Optical mapping. The finished genome (5.3 Mb chromosome and two plasmids pXcaw19 and pXcaw58) of Xcaw was annotated, curated and compared with XccA genome. Protein blast revealed multiple genes including type III secretion system (TIIISS) effectors xopAF and xopAG are present in Xcaw but absent in XccA. Comparative genomic analysis showed various changes in genes encoding LPS and type IV secretion system. Furthermore, RNA-Seq was used to compare expression profile of Xcaw and XccA in nutrient rich (NB) medium and XVM2 medium which is known to mimic the intercellular space of plant cells using Illumina sequencing. Multiple avirulence/effector genes were over-expressed in Xcaw compared to XccA which might contribute to the limited host range of Xcaw compared to XccA. The overexpression of genes involved in cell wall degradation, attachment, ROS (reactive oxygen species) scavenging, nutrient transportation in XccA might contribute to its expanding of host range. Our data suggest that both gene content and gene expression contribute to difference in virulence and host range of bacterial pathogens. This study lays the foundation to further characterize the mechanisms for virulence and host range of strains of X. citri subsp. citri and other bacterial pathogens. mRNA expression profiles of Xcc strain A and Aw were generated in 2 media: NB and XVM2 by deep sequencing, in triplicate, using Illumina GAII.

Project description:Xanthomonas is one important model microbe to study the molecular determinants of virulence and host range of pathogens since Xanthomonas is capable of infecting numerous monocotyledonous and dicotyledonous plants. Among the plant diseases caused by Xanthomonas, X. citri subsp. citri (Xcc) causes citrus canker, which has significant impact on citrus production. Xcc is classified into different strains primarily by host range including A and Aw. The A (Asiatic) strain (XccA) has a wide host range and is most virulent, whereas Aw (Wellington) strain has restricted host range including Key or Mexican lime and alemow. We hypothesized that not only gene content but also gene expression contributes to the difference in virulence and host range of closely related strains. To test our hypothesis, comparative genomic and transcriptome analyses were conducted to study the two closely related Xcc A and Aw strains. The genome of X. citri subsp. citri strain Aw12879 (Xcaw) was completely sequenced using 454 Pyrosequencing, Illumina sequencing and Optical mapping. The finished genome (5.3 Mb chromosome and two plasmids pXcaw19 and pXcaw58) of Xcaw was annotated, curated and compared with XccA genome. Protein blast revealed multiple genes including type III secretion system (TIIISS) effectors xopAF and xopAG are present in Xcaw but absent in XccA. Comparative genomic analysis showed various changes in genes encoding LPS and type IV secretion system. Furthermore, RNA-Seq was used to compare expression profile of Xcaw and XccA in nutrient rich (NB) medium and XVM2 medium which is known to mimic the intercellular space of plant cells using Illumina sequencing. Multiple avirulence/effector genes were over-expressed in Xcaw compared to XccA which might contribute to the limited host range of Xcaw compared to XccA. The overexpression of genes involved in cell wall degradation, attachment, ROS (reactive oxygen species) scavenging, nutrient transportation in XccA might contribute to its expanding of host range. Our data suggest that both gene content and gene expression contribute to difference in virulence and host range of bacterial pathogens. This study lays the foundation to further characterize the mechanisms for virulence and host range of strains of X. citri subsp. citri and other bacterial pathogens.

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