HrpG and HrpX play global roles in coordinating different virulence traits of Xanthomonas axonopodis pv. citri

Xanthomonas axonopodis pv. citri is the causal agent of citrus canker, which is one of the most serious diseases of citrus. To understand the virulence mechanisms of X. axonopodis pv. citri, we designed and conducted genome-wide microarray analyses to characterize the HrpG and HrpX regulons, which are critical for the pathogenicity of X. axonopodis pv. citri. Our analyses revealed that 232 and 181 genes belonged to the HrpG and HrpX regulons, respectively. In total, 123 genes were overlapped in the two regulons at any of the three selected timepoints representing three growth stages of X. axonopodis pv. citri in XVM2 medium. Our results showed that HrpG and HrpX regulated all 24 type III secretion system genes, 23 type III secretion system effector genes, and 29 type II secretion system substrate genes. Our data revealed that X. axonopodis pv. citri 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; and phenolic catabolism, through HrpX and HrpG. We found that 124 and 90 unknown genes were controlled by HrpG and HrpX, respectively. Our results suggest that HrpG and HrpX interplay with a global signaling network and co-ordinate the expression of multiple virulence factors for modification and adaption of host environment during X. axonopodis pv. citri infection.     

Genomic Relatedness of Xanthomonas campestris Strains Causing Diseases of Citrus

Xanthomonas campestris strains that cause disease in citrus were compared by restriction endonuclease analysis of DNA fragments separated by pulsed-field gel electrophoresis and by DNA reassociation. Strains of X. campestris pv. citrumelo, which cause citrus bacterial spot, were, on average, 88% related to each other by DNA reassociation, although these strains exhibited diverse restriction digest patterns. In contrast, strains of X. campestris pv. citri groups A and B, which cause canker A and canker B, respectively, had relatively homogeneous restriction digest patterns. The groups of strains causing these three different citrus diseases were examined by DNA reassociation and were found to be from 55 to 63% related to one another. Several pathovars of X. campestris, previously shown to cause weakly aggressive symptoms on citrus, ranged from 83 to 90% similar to X. campestris pv. citrumelo by DNA reassociation. The type strain of X. campestris pv. campestris ranged from 30 to 40% similar in DNA reassociation experiments to strains of X. campestris pv. citrumelo and X. campestris pv. citri groups A and B. Whereas DNA reassociation quantified the difference between relatively unrelated groups of bacterial strains, restriction endonuclease analysis distinguished between closely related strains.     

Insights into genome plasticity and pathogenicity of the plant pathogenic bacterium Xanthomonas campestris pv. vesicatoria revealed by the complete genome sequence

The gram-negative plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria is the causative agent of bacterial spot disease in pepper and tomato plants, which leads to economically important yield losses. This pathosystem has become a well-established model for studying bacterial infection strategies. Here, we present the whole-genome sequence of the pepper-pathogenic Xanthomonas campestris pv. vesicatoria strain 85-10, which comprises a 5.17-Mb circular chromosome and four plasmids. The genome has a high G+C content (64.75%) and signatures of extensive genome plasticity. Whole-genome comparisons revealed a gene order similar to both Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris and a structure completely different from Xanthomonas oryzae pv. oryzae. A total of 548 coding sequences (12.2%) are unique to X. campestris pv. vesicatoria. In addition to a type III secretion system, which is essential for pathogenicity, the genome of strain 85-10 encodes all other types of protein secretion systems described so far in gram-negative bacteria. Remarkably, one of the putative type IV secretion systems encoded on the largest plasmid is similar to the Icm/Dot systems of the human pathogens Legionella pneumophila and Coxiella burnetii. Comparisons with other completely sequenced plant pathogens predicted six novel type III effector proteins and several other virulence factors, including adhesins, cell wall-degrading enzymes, and extracellular polysaccharides.     

Functional analysis of the early chlorosis factor gene

Chlorosis is one of the symptoms of bacterial spot disease caused by Xanthomonas campestris pv. vesicatoria, which induces chlorosis before any other symptoms appear on tomato. We report characterization of a 2.1-kb gene called early chlorosis factor (ecf). The gene ecf encodes a hydrophobic protein with similarity to four other proteins in plant pathogens, including HolPsyAE, and uncharacterized gene products from X. campestris pv. campestris and X. axonopodis pv. citri, and, at the tertiary structure level, to colicin Ia from Escherichia coli. We demonstrate that the associated phenotype is hrp dependent, and that the ecf gene product appears to be translocated to host cells. The gene ecf has no impact on electrolyte leakage or on bacterial growth in planta in response to infection. Concentrated culture filtrates do not produce chlorosis. Study of its role in Xanthomonas spp.-tomato interactions will forward our understanding of symptom production by plant pathogens and allows further investigation into the mechanisms of bacterial virulence and production of symptoms.     

Sensitive and specific detection of Xanthomonas axonopodis pv. citri by PCR using pathovar specific primers based on hrpW gene sequences

A sensitive and specific assay was developed to detect citrus bacterial canker caused by Xanthomonas axonopodis pv. citri, in leaves and fruits of citrus. Primers XACF and XACR from hrpW homologous to pectate lyase, modifying the structure of pectin in plants, were used to amplify a 561 bp DNA fragment. PCR technique was applied to detect the pathogen in naturally or artificially infected leaves of citrus. The PCR product was only produced from X. axonopodis pv. citri among 26 isolates of Xanthomonas strains, Escherichia coli (O157:H7), Pectobacterium carotovorum subsp. carotovorum, and other reference microbes.     

A LOV protein modulates the physiological attributes of Xanthomonas axonopodis pv. citri relevant for host plant colonization

Recent studies have demonstrated that an appropriate light environment is required for the establishment of efficient vegetal resistance responses in several plant-pathogen interactions. The photoreceptors implicated in such responses are mainly those belonging to the phytochrome family. Data obtained from bacterial genome sequences revealed the presence of photosensory proteins of the BLUF (Blue Light sensing Using FAD), LOV (Light, Oxygen, Voltage) and phytochrome families with no known functions. Xanthomonas axonopodis pv. citri is a Gram-negative bacterium responsible for citrus canker. The in silico analysis of the X. axonopodis pv. citri genome sequence revealed the presence of a gene encoding a putative LOV photoreceptor, in addition to two genes encoding BLUF proteins. This suggests that blue light sensing could play a role in X. axonopodis pv. citri physiology. We obtained the recombinant Xac-LOV protein by expression in Escherichia coli and performed a spectroscopic analysis of the purified protein, which demonstrated that it has a canonical LOV photochemistry. We also constructed a mutant strain of X. axonopodis pv. citri lacking the LOV protein and found that the loss of this protein altered bacterial motility, exopolysaccharide production and biofilm formation. Moreover, we observed that the adhesion of the mutant strain to abiotic and biotic surfaces was significantly diminished compared to the wild-type. Finally, inoculation of orange (Citrus sinensis) leaves with the mutant strain of X. axonopodis pv. citri resulted in marked differences in the development of symptoms in plant tissues relative to the wild-type, suggesting a role for the Xac-LOV protein in the pathogenic process. Altogether, these results suggest the novel involvement of a photosensory system in the regulation of physiological attributes of a phytopathogenic bacterium. A functional blue light receptor in Xanthomonas spp. has been described for the first time, showing an important role in virulence during citrus canker disease.     

Characterization of the hrpF pathogenicity peninsula of Xanthomonas oryzae pv. oryzae

The hrp gene cluster of Xanthomonas spp. contains genes for the assembly and function of a type III secretion system (TTSS). The hrpF genes reside in a region between hpaB and the right end of the hrp cluster. The region of the hrpF gene of Xanthomonas oryzae pv. oryzae is bounded by two IS elements and also contains a homolog of hpaF of X. campestris pv. vesicatoria and two newly identified genes, hpa3 and hpa4. A comparison of the hrp gene clusters of different species of Xanthomonas revealed that the hrpF region is a constant yet more variable peninsula of the hrp pathogenicity island. Mutations in hpaF, hpa3, and hpa4 had no effect on virulence, whereas hrpF mutants were severely reduced in virulence on susceptible rice cultivars. The hrpF genes from X. campestris pv. vesicatoria, X. campestris pv. campestris, and X. axonopodis pv. citri each were capable of restoring virulence to the hrpF mutant of X. oryzae pv. oryzae. Correspondingly, none of the Xanthomonas pathovars with hrpF from X. oryzae pv. oryzae elicited a hypersensitive reaction in their respective hosts. Therefore, no evidence was found for hrpF as a host-specialization factor. In contrast to the loss of Bs3-dependent reactions by hrpF mutants of X. campestris pv. vesicatoria, hrpF mutants of X. oryzae pv. oryzae with either avrXa10 or avrXa7 elicited hypersensitive reactions in rice cultivars with the corresponding R genes. A double hrpFxoo-hpa1 mutant also elicited an Xa10-dependent resistance reaction. Thus, loss of hrpF, hpal, or both may reduce delivery or effectiveness of type III effectors. However, the mutations did not completely prevent the delivery of effectors from X. oryzae pv. oryzae into the host cells.     

In vivo proteome analysis of Xanthomonas campestris pv. campestris in the interaction with the host plant Brassica oleracea

The genus Xanthomonas is composed of several species that cause severe crop losses around the world. In Latin America, one of the most relevant species is Xanthomonas campestris pv. campestris, which is responsible for black rot in cruciferous plants. This pathogen causes yield losses in several cultures, including cabbage, cauliflower and broccoli. Although the complete structural genome of X. campestris pv. campestris has been elucidated, little is known about the protein expression of this pathogen in close interaction with the host plant. Recently, a method for in vivo analysis of Xanthomonas axonopodis pv. citri was developed. In the present study, this technique was employed for the characterization of the protein expression of X. campestris pv. campestris in close interaction with the host plant Brassica oleracea. The bacterium was infiltrated into leaves of the susceptible cultivar and later recovered for proteome analysis. Recovered cells were used for protein extraction and separated by two-dimensional electrophoresis. Proteins were analysed by peptide mass fingerprinting or de novo sequencing and identified by searches in public databases. The approach used in this study may be extremely useful in further analyses in order to develop novel strategies to control this important plant pathogen.     

Virulence deficiency caused by a transposon insertion in the purH gene of Xanthomonas oryzae pv. oryzae

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. We have identified a Tn5-induced virulence-deficient mutant (BXO1704) of X. oryzae pv. oryzae. The BXO1704 mutant exhibited growth deficiency in minimal medium but was proficient in inducing a hypersensitive response in a non-host tomato plant. Sequence analysis of the chromosomal DNA flanking the Tn5 insertion indicated that the Tn5 insertion is in the purH gene, which is highly homologous to purH genes of other closely related plant pathogenic bacteria Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris. Purine supplementation reversed the growth deficiency of BXO1704 in minimal medium. These results suggest that the virulence deficiency of BXO1704 may be due to the inability to use sufficient purine in the host.     

Detection of Xanthomonas campestris pv. citri by the polymerase chain reaction method.

pFL1 is a pUC9 derivative that contains a 572-bp EcoRI insert cloned from plasmid DNA of Xanthomonas campestris pv. citri XC62. The nucleotide sequence of pFL1 was determined, and the sequence information was used to design primers for application of the polymerase chain reaction (PCR) to the detection of X. campestris pv. citri, the causal agent of citrus bacterial canker disease. Seven 18-bp oligonucleotide primers were designed and tested with DNA from X. campestris pv. citri strains and other strains of X. campestris associated with Citrus spp. as templates in the PCR. Four primer pairs directed the amplification of target DNA from X. campestris pv. citri strains but not from strains of X. campestris associated with a different disease, citrus bacterial spot. Primer pair 2-3 directed the specific amplification of target DNA from pathotype A but not other pathotypes of X. campestris pv. citri. A pH 9.0 buffer that contained 1% Triton X-100 and 0.1% gelatin was absolutely required for the successful amplification of the target DNA, which was 61% G+C. Limits of detection after amplification and gel electrophoresis were 25 pg of purified target DNA and about 10 cells when Southern blots were made after gel electrophoresis and probed with biotinylated pFL1. This level of detection represents an increase in sensitivity of about 100-fold over that of dot blotting with the same hybridization probe. PCR products of the expected sizes were amplified from DNA extracted from 7-month-old lesions from which viable bacteria could not be isolated. These products were confirmed to be specific for X. campestris pv. citri by Southern blotting. This PCR-based detection protocol will be a useful addition to current methods of detection of this pathogen, which is currently the target of international quarantine measures.     

Biofilm formation, epiphytic fitness, and canker development in Xanthomonas axonopodis pv. citri

The phytopathogenic bacterium Xanthomonas axonopodis pv. citri is responsible for the canker disease affecting citrus plants throughout the world. Here, we have evaluated the role of bacterial attachment and biofilm formation in leaf colonization during canker development on lemon leaves. Crystal violet staining and confocal laser scanning microscopy analysis of X. axonopodis pv. citri strains expressing the green fluorescent protein were used to evaluate attachment and biofilm formation on abiotic and biotic (leaf) surfaces. Wild-type X. axonopodis pv. citri attached to and formed a complex, structured biofilm on glass in minimal medium containing glucose. Similar attachment and structured biofilm formation also were seen on lemon leaves. An X. axonopodis pv. citri gumB mutant strain, defective in production of the extracellular polysaccharide xanthan, did not form a structured biofilm on either abiotic or biotic surfaces. In addition, the X. axonopodis pv. citri gumB showed reduced growth and survival on leaf surfaces and reduced disease symptoms. These findings suggest an important role for formation of biofilms in the epiphytic survival of X. axonopodis pv. citri prior to development of canker disease.     

Site-directed gene replacement of the phytopathogen Xanthomonas axonopodis pv. citri

In this work we defined experimental conditions for site-directed gene replacement of the Xanthomonas axonopodis pv. citri (Xac), an economically relevant pathogen of citrus plants. The procedure involved, first, optimizing the electrotransformation conditions of the Xac 306 strain and, second, constructing non-replicative suicide vectors carrying knockout copies of the target gene. Using specific experimental conditions, transformation efficiencies of Xac were at least 100 fold higher than those achieved with electroporation protocols previously designed for X. campestris transformation. Successful gene replacement events were achieved with a suicide vector derived from R6K plasmid (pWR-SS) but not with those with ColE1 replication origin. We have chosen the oppA as a target gene, encoding the binding component (OppA) of the major oligopeptide uptake system found in the genome of the Xac 306 strain, although not in X. campestris pv. campestris (Xcc). Defining the experimental conditions, which allow for the specific mutagenesis of the Xac 306 strain, represents a step in the understanding of both genetics and physiology of this economically important bacterial species.     

Evaluation of the Biolog Substrate Utilization System To Identify and Assess Metabolic Variation among Strains of Xanthomonas campestris pv. Citri

Metabolic fingerprints of 148 strains of Xanthomonas campestris pv. citri originating from 24 countries and associated with various forms of citrus bacterial canker disease (CBCD) were obtained by using the Biolog substrate utilization system. Metabolic profiles were used to attempt strain identification. Only 6.8% of the studied strains were correctly identified when the commercial Microlog 2N data base was used alone. When the data base was supplemented with data from 54 strains of X. campestris pv. citri (40 CBCD-A strains, 8 CBCD-B strains, and 6 CBCD-C strains) and data from 43 strains of X. campestris associated with citrus bacterial spot disease, the percentage of correct identifications was 70%. Thus, it is recommended that users supplement the commercial data base with additional data prior to using the program for identification purposes. The utilization of Tween 40 in conjunction with other tests can help to differentiate strains associated with CBCD and citrus bacterial spot disease. These results confirmed the separation of X. campestris pv. citri into different subgroups (strains associated with Asiatic citrus canker [CBCD-A], cancrosis B [CBCD-B], and Mexican lime canker [CBCD-C]). The utilization of l-fucose, d-galactose, and alaninamide can be used as markers to differentiate strains associated with these groups. A single strain associated with bacteriosis of Mexican lime in Mexico (CBCD-D) was closely similar to CBCD-B strains.     

Reduced genetic variation occurs among genes of the highly clonal plant pathogen Xanthomonas axonopodis pv. vesicatoria, including the effector gene avrBs2

The bacterial plant pathogen Xanthomonas axonopodis pv. vesicatoria, also known as Xanthomonas campestris pv. vesicatoria group A, is the causal agent of bacterial spot in pepper and tomato. In order to test different models that may explain the coevolution of avrBs2 with its host plants, we sequenced avrBs2 and six chromosomal loci (total of 5.5 kb per strain) from a global sample of 55 X. axonopodis pv. vesicatoria strains collected from diseased peppers. We found an extreme lack of genetic variation among all X. axonopodis pv. vesicatoria genomic loci (average nucleotide diversity, pi = 9.1 x 10(-5)), including avrBs2. This lack of diversity is consistent with X. axonopodis pv. vesicatoria having undergone a recent population bottleneck and/or selective sweep followed by population expansion. Coalescent analysis determined that approximately 1.4 x 10(4) to 7.16 x 10(4) bacterial generations have passed since the most recent common ancestor (MRCA) of the current X. axonopodis pv. vesicatoria population. Assuming a range of 50 to 500 bacterial generations per year, only 28 to 1,432 years have passed since the MRCA. This time frame coincides with human intervention with the pathogen's host plants, from domestication to modern agricultural practices. Examination of 19 mutated (loss-of-function) avrBs2 alleles detected nine classes of mutations. All mutations affected protein coding, while no synonymous changes were found. The nature of at least one of the avrBs2 mutations suggests that it may be possible to observe one stage of an evolutionary arms race as X. axonopodis pv. vesicatoria responds to selection pressure to alter avrBs2 to escape host plant resistance.     

PhyA, a secreted protein of Xanthomonas oryzae pv. oryzae, is required for optimum virulence and growth on phytic acid as a sole phosphate source

Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, a serious disease of rice. We have identified a novel virulence deficient mutant (BXO1691) of X. oryzae pv. oryzae that has a Tn5 insertion in an open reading frame (phyA; putative phytase A) encoding a 373-amino acid (aa) protein containing a 28-aa predicted signal peptide. Extracellular protein profiles revealed that a 38-kDa band is absent in phyA mutants as compared with phyA+ strains. A BLAST search with phyA and its deduced polypeptide sequence indicated significant similarity with conserved hypothetical proteins in Xanthomonas axonopodis pv. citri and Xanthomonas campestris pv. campestris and limited homology to secreted phytases of Bacillus species. Homology modeling with a Bacillus phytase as the template suggests that the PhyA protein has a similar six-bladed beta-propeller architecture and exhibits conservation of certain critical active site residues. Phytases are enzymes that are involved in degradation of phytic acid (inositol hexaphosphate), a stored form of phosphate in plants. The phyA mutants exhibit a growth deficiency in media containing phytic acid as a sole phosphate source. Exogenous phosphate supplementation promotes migration of phyA X. oryzae pv. oryzae mutants in rice leaves. These results suggest that the virulence deficiency of phyA mutants is, at least in part, due to inability to use host phytic acid as a source of phosphate. phyA-like genes have not been previously reported to be involved in the virulence of any plant pathogenic bacterium.     

Mutation in the xpsD gene of Xanthomonas axonopodis pv. citri affects cellulose degradation and virulence

The Gram-negative bacterium Xanthomonas axonopodis pv. citri, the causal agent of citrus canker, is a major threat to the citrus industry worldwide. Although this is a leaf spot pathogen, it bears genes highly related to degradation of plant cell walls, which are typically found in plant pathogens that cause symptoms of tissue maceration. Little is known on Xac capacity to cause disease and hydrolyze cellulose. We investigated the contribution of various open reading frames on degradation of a cellulose compound by means of a global mutational assay to selectively screen for a defect in carboxymethyl cellulase (CMCase) secretion in X. axonopodis pv. citri. Screening on CMC agar revealed one mutant clone defective in extracellular glycanase activity, out of nearly 3,000 clones. The insertion was located in the xpsD gene, a component of the type II secretion system (T2SS) showing an influence in the ability of Xac to colonize tissues and hydrolyze cellulose. In summary, these data show for the first time, that X. axonopodis pv. citri is capable of hydrolyzing cellulose in a T2SS-dependent process. Furthermore, it was demonstrated that the ability to degrade cellulose contributes to the infection process as a whole.     

四种黄单胞菌的基因芯片检测方法的建立

结合双重PCR和基因芯片技术同时检测和鉴定我国检疫性细菌,包括水稻白叶枯病菌(Xanthomonas oryzae pv.oryzae,Xoo)、水稻细菌性条斑病菌(X.oryzae pv.oryzicola,Xooc)、柑桔溃疡病菌(X.axonopodis pv.citri,Xac)以及严重危害十字花科作物的甘蓝黑腐病菌(Xanthomonas campestris pv.campestris,Xcc)。以铁载体受体(Putative siderophore receptor)基因序列和RNA多聚酶西格玛因子(RNA polymerase sigma factor,rpoD)基因序列为靶标,设计引物和特异性探针能够同时检测这4种重要的病原菌。对17个细菌菌株进行芯片检测,仅4种靶标菌得到阳性结果,证明此方法具有很高的特异性。4种致病菌基因组DNA的检测灵敏度约为3 pg。检测结果表明,建立的基因芯片检测方法特异性强,能实现上述4种黄单胞菌的准确检测和鉴定,具有良好的应用前景。     

Comparative analyses of Xanthomonas and Xylella complete genomes

Computational analyses of four bacterial genomes of the Xanthomonadaceae family reveal new unique genes that may be involved in adaptation, pathogenicity, and host specificity. The Xanthomonas genus presents 3636 unique genes distributed in 1470 families, while Xylella genus presents 1026 unique genes distributed in 375 families. Among Xanthomonas-specific genes, we highlight a large number of cell wall degrading enzymes, proteases, and iron receptors, a set of energy metabolism genes, second copy of the type II secretion system, type III secretion system, flagella and chemotactic machinery, and the xanthomonadin synthesis gene cluster. Important genes unique to the Xylella genus are an additional copy of a type IV pili gene cluster and the complete machinery of colicin V synthesis and secretion. Intersections of gene sets from both genera reveal a cluster of genes homologous to Salmonella's SPI-7 island in Xanthomonas axonopodis pv citri and Xylella fastidiosa 9a5c, which might be involved in host specificity. Each genome also presents important unique genes, such as an HMS cluster, the kdgT gene, and O-antigen in Xanthomonas axonopodis pv citri; a number of avrBS genes and a distinct O-antigen in Xanthomonas campestris pv campestris, a type I restriction-modification system and a nickase gene in Xylella fastidiosa 9a5c, and a type II restriction-modification system and two genes related to peptidoglycan biosynthesis in Xylella fastidiosa temecula 1. All these differences imply a considerable number of gene gains and losses during the divergence of the four lineages, and are associated with structural genome modifications that may have a direct relation with the mode of transmission, adaptation to specific environments and pathogenicity of each organism.     

Laterally transferred genomic islands in Xanthomonadales related to pathogenicity and primary metabolism.

Lateral gene transfer (LGT) is considered as one of the drivers in bacterial genome evolution, usually associated with increased fitness and/or changes in behavior, especially if one considers pathogenic vs. non-pathogenic bacterial groups. The genomes of two phytopathogens, Xanthomonas campestris pv. campestris and Xanthomonas axonopodis pv. citri, were previously inspected for genome islands originating from LGT events, and, in this work, potentially early and late LGT events were identified according to their altered nucleotide composition. The biological role of the islands was also assessed, and pathogenicity, virulence and secondary metabolism pathways were functions highly represented, especially in islands that were found to be recently transferred. However, old islands are composed of a high proportion of genes related to cell primary metabolic functions. These old islands, normally undetected by traditional atypical composition analysis, but confirmed as product of LGT by atypical phylogenetic reconstruction, reveal the role of LGT events by replacing core metabolic genes normally inherited by vertical processes.     

Non-gamma-proteobacteria gene islands contribute to the Xanthomonas genome

Horizontal gene transfer, a process through which genomes acquire sequences from distantly related organisms, is believed to be a major source of genetic diversity in bacteria. A central question concerning the impact of gene transfer on bacterial genome evolution is the proportion of horizontally transferred sequences within genomes. Through BLAST search, we found that the genomes of two phytopathogens, Xanthomonas campestris pv. campestris and Xanthomonas axonopodis pv. citri, have close to 40% of the genes with the highest similarity to genes from phylogenetically distant organisms (non-gamma-proteobacteria). Most of these genes are found to be contiguous in the genome, forming genome islands, which may have been transferred from other organisms. Overall, the total number of genes within genome islands corresponds to almost one quarter of the entire xanthomonad genomes. Interestingly, many of the genes in these islands are functionally related to plant pathogenesis and virulence. Thus, these results suggest that horizontally transferred genes are clustered in the genome, and may facilitate fitness in new environments, as in the case of plant-bacteria interaction.