The Genome Sequencing And Comparative Genomics Analysis Of Xanthomonas Axonopodis Pv. Citri

Citrus canker caused by Xanthomonas axonopodis pv. citri(Xac) is one of the most severe bacterial disease of citrus that threatened to the citrus industry worldwide. The disease can affect the plant growing and influence the fruit quality. With the rapid development of DNA sequencing technology, genomics research has gradually become a new research hotspot. Although the various study of Xac has been underway for many, there is still more space need to be improve on the molecular mechanism of Xac. In order to study host range and pathogenic mechanism between Xac A and Xac A~W, the Xac A strain isolated from the infected orange in Jiangxi and sequencing through Illumina Miseq sequencing platform. Further comparative genome analysis were conducted. The main research result was described as follows: 1. Genome sequencing of Xac strain and analysisIn this study, the whole genome of jx-6 strain was sequenced by using Illumina Miseq sequencing platform. The jx-6 strain, which consists of a circular chromosome(Gen Bank accession number: CP011827.1) with the genome size of 5,124,792 bp and two plasmids(Gen Bank accession number: CP013664.1 and CP013665.1). The genome contains 4357 protein-coding genes and 54 t RNA genes and six 16S-23S-5S ribosomal r RNA operon. The genome GC content of jx-6 genome is 64.8%. Moreover, the basic characteristics of inserted sequences, secretion system, gene island, phage and CRISPR of jx-6 genome are predicted. The result show that the genome of jx-6 strain has 14 crdible IS elements, 42 GIs,3 candidate prophage regions and 5 CRISPR loci. 2. Comparative genomic analysis of Xac strainsThe comparative genomic analysis were performed among different Xac strains, including three Xac A(jx-6, gd3, 306) and two Xac A~W strains(A~W13, A~W12879) were selected and analyzed. Orthologous analysis showed that there are 3887 orthologous were determined as the core gene clusters that shared by all 5 strains. Only Xac A~W have 216 specific gene cluster, which are associated as biological metabolism and cellular metabolic processes. The whole genomic nucleotide sequence collinearity comparative analysis results showed that there were overturn, translocation and inversion among the genomes of Xac A~W. In addition, the prophage prediction analysis of Xac A and Xac A~W show that Xac A contains three prophage regions, while Xac A~W contains five prophage regions.We found the prophage2 in Xac A and prophage2 in Xac A~W belong to the same kind of phage. Prophage4 and prophage5 are just inserted in the low levels of linear parts between Xac A and Xac A~W. These prophage insertion may be caused the host specialization of Xac A~W. 3. The analysis of the CRISPR/cas system of Xac strainsClustered regularly interspaced short palindromic repeats(CRISPR), which provides effective resistance against foreign nucleic-acid elements, for instance, the phage and plasmid. Analysis of the strain jx-6 indicated that five candidate CRISPR loci in the jx-6 genome and the CIRSPR associated genes(cas genes) had been found at upstream area of CRISPR5 locus. The comparison of CRISPR 5 locus on different Xac strains showed the repeat unit of CRISPR 5 locus are variable that can be hypothesised the CRISPR 5 could be activity. In this study, to confirm the variable of repeat unit on CRISPR 5, Xac strains from Guangdong, Guangxi, Jiangxi were isolated and amplified through PCR and the amplicon was sequenced. The result is consistent with the hypothesis that indicated the CRISPR 5 could be active. 4. The phylogenetic relationship analysis of Xac strainsIn this study, the phylogenetic relationship was predicted by series connection of Xac seven housekeeping genes(atp D,dna K,efp,fyu A,gln A,gyr B,rpo D). Strains in the phylogenetic tree including 22 strains of Xac, 17 strains of Xac A and 5 strains of Xac A~W genome sequences. The phylogenetic tree shows that all Xac A strains can be grouped together, while all Xac Aw strains were in the same branch. The jx-6 strain is close to strain BL18. Interestingly, the above results are also consistent with the COG results in the genomics analysis and orthologous analysis results in the comparative genomics.