Project description:Quorum sensing (QS) in Xanthomonas axonopodis pv. citri, the causal agent of citrus canker, is mediated by a diffusible signal factor (DSF). QS is required for the full virulence of X. axonopodis pv. citri in planta. Mutations in rpfF, rpfC and rpfG, the core genes of QS, decreased the production of extracellular proteases and bacterial motility. Comparison of the transcriptomes of QS mutants with that of the wild type stain revealed that QS temporally regulates the expression of a large set of genes, including genes involved in chemotaxis and flagellar biosynthesis, genes related to metabolism, genes encoding virulence traits such as type II secretion system substates, type III secretion system and effectors. Cross talk between the QS regulon and the HrpG regulon has also been identified by 62 common genes shared by both regulons. The temporal regulation of the QS regulon and cross talk with the HrpG regulon suggest the important role of QS in citrus canker infection, including attachment, invasion and growth in host apoplast.
Project description:Quorum sensing (QS) in Xanthomonas axonopodis pv. citri, the causal agent of citrus canker, is mediated by a diffusible signal factor (DSF). QS is required for the full virulence of X. axonopodis pv. citri in planta. Mutations in rpfF, rpfC and rpfG, the core genes of QS, decreased the production of extracellular proteases and bacterial motility. Comparison of the transcriptomes of QS mutants with that of the wild type stain revealed that QS temporally regulates the expression of a large set of genes, including genes involved in chemotaxis and flagellar biosynthesis, genes related to metabolism, genes encoding virulence traits such as type II secretion system substates, type III secretion system and effectors. Cross talk between the QS regulon and the HrpG regulon has also been identified by 62 common genes shared by both regulons. The temporal regulation of the QS regulon and cross talk with the HrpG regulon suggest the important role of QS in citrus canker infection, including attachment, invasion and growth in host apoplast. Three mutants and two time-points experiment. Four biological and dye-swap replicates of each strain per each time-point were used. In total, there were two datasets of the rpfF mutant: 1) rpfF mutant vs. wild type strain at 11 h, and 2) rpfF mutant vs. wild type strain at 25 h; two datasets of the rpfC mutant: 1) rpfC mutant vs. wild type strain at 11 h, and 2) rpfC mutant vs. wild type strain at 25 h; and two datasets of the rpfG mutant: 1) rpfG mutant vs. wild type strain at 11 h, and 2) rpfG mutant vs. wild type strain at 25 h.
Project description:In this project, we have extracted the flagellum of the citrus canker pathogens Xanthomonas citri and Xanthomonas aurantifolii pathotype C and the proteins associated with these preparations were resolved by SDS-PAGE and identified by mass spectrometry. The major 50 kDa band, observed in both samples, correspond to the flagellin C (FliC) proteins. All the peptides identified by mass spectrometry shown here showed perfect matches to the X. citri and X. aurantifolii FliC proteins deposited in the GenBank (NCBI reference sequences WP_003482972 and WP_007962409, respectively).
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.
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 axonopodis pv. citri (XAC) is the causal agent of citrus canker which is one of the most serious diseases of citrus. To understand the virulence mechanism of XAC, we designed and conducted genome-wide microarray analyses to characterize the regulons of HrpG and HrpX, which are critical for the pathogenicity of XAC. Our analyses revealed that the expression of 232 genes and 181 genes belonged to the HrpG and HrpX regulons, respectively. Totally, 123 genes were overlapped in the two regulons at any of the three time-points. Our results showed that HrpG and HrpX regulated all 25 T3SS genes, 22 T3SS effector genes, and 29 T2SS substrate genes. Our data showed that XAC regulates multiple cellular activities responding to the host environment, such as amino acid biosynthesis, oxidative phosphorylation, pentose-phosphate pathway, transport of sugar, iron and potassium, the phenolic compounds catabolism pathway (pcaQ) through HrpX and HrpG. We also identified 124 and 90 unknown genes controlled by HrpG and HrpX, respectively. Our data suggest that a cross-talk exists between HrpG and quorum sensing system in XAC. We also identified genes involved in chemotaxis and flagellar biosynthesis, production of extracellular enzymes, metabolic pathways, as well as signal transduction controlled by HrpG/HrpX. Our results suggest that HrpG and HrpX interplay with global signaling network and co-ordinate the expression of multiple virulence factors for modification and adaption of host environment during XAC infection.
Project description:Xanthomonas axonopodis pv. citri (XAC) is the causal agent of citrus canker which is one of the most serious diseases of citrus. To understand the virulence mechanism of XAC, we designed and conducted genome-wide microarray analyses to characterize the regulons of HrpG and HrpX, which are critical for the pathogenicity of XAC. Our analyses revealed that the expression of 232 genes and 181 genes belonged to the HrpG and HrpX regulons, respectively. Totally, 123 genes were overlapped in the two regulons at any of the three time-points. Our results showed that HrpG and HrpX regulated all 25 T3SS genes, 22 T3SS effector genes, and 29 T2SS substrate genes. Our data showed that XAC regulates multiple cellular activities responding to the host environment, such as amino acid biosynthesis, oxidative phosphorylation, pentose-phosphate pathway, transport of sugar, iron and potassium, the phenolic compounds catabolism pathway (pcaQ) through HrpX and HrpG. We also identified 124 and 90 unknown genes controlled by HrpG and HrpX, respectively. Our data suggest that a cross-talk exists between HrpG and quorum sensing system in XAC. We also identified genes involved in chemotaxis and flagellar biosynthesis, production of extracellular enzymes, metabolic pathways, as well as signal transduction controlled by HrpG/HrpX. Our results suggest that HrpG and HrpX interplay with global signaling network and co-ordinate the expression of multiple virulence factors for modification and adaption of host environment during XAC infection. Two mutants and three time-points experiment. Four biological and dye-swap replicates of each strain per each time-point were used. Totally, there were three datasets of hrpG mutant: 1) hrpG mutant vs. wild type strain at time-point A; 2) hrpG mutant vs. wild type strain at time-point B; 3) hrpG mutant vs. wild type strain at time-point C. There are also three datasets of hrpX mutant: 1) hrpX mutant vs. wild type strain at time-point A; 2) hrpX mutant vs. wild type strain at time-point B; 3) hrpX mutant vs. wild type strain at time-point C.