Novel insights into the genomic basis of citrus canker based on the genome sequences of two strains of Xanthomonas fuscans subsp. aurantifolii

Background: Citrus canker is a disease that has severe economic impact on the citrus industry worldwide. There are three types of canker, called A, B, and C. The three types have different phenotypes and affect different citrus species. The causative agent for type A is Xanthomonas citri subsp. citri, whose genome sequence was made available in 2002. Xanthomonas fuscans subsp. aurantifolii strain B causes canker B and Xanthomonas fuscans subsp. aurantifolii strain C causes canker C. Results: We have sequenced the genomes of strains B and C to draft status. We have compared their genomic content to X. citri subsp. citri and to other Xanthomonas genomes, with special emphasis on type III secreted effector repertoires. In addition to pthA, already known to be present in all three citrus canker strains, two additional effector genes, xopE3 and xopAI, are also present in all three strains and are both located on the same putative genomic island. These two effector genes, along with one other effector-like gene in the same region, are thus good candidates for being pathogenicity factors on citrus. Numerous gene content differences also exist between the three cankers strains, which can be correlated with their different virulence and host range. Particular attention was placed on the analysis of genes involved in biofilm formation and quorum sensing, type IV secretion, flagellum synthesis and motility, lipopolysacharide synthesis, and on the gene xacPNP, which codes for a natriuretic protein. Conclusion: We have uncovered numerous commonalities and differences in gene content between the genomes of the pathogenic agents causing citrus canker A, B, and C and other Xanthomonas genomes. Molecular genetics can now be employed to determine the role of these genes in plant-microbe interactions. The gained knowledge will be instrumental for improving citrus canker control.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho Nacional de Desenvolvimento CientIfico e Tecnologico (CNPq)Coordenacao para Aperfeicoamento de Pessoal de Ensino Superior (CAPES)Fundo de Defesa da Citricultura (FUNDECITRUS

Microbial Community Composition of ZC1 and ZC2 metagenomes.

<p>Unassembled reads annotated on MG-RAST were analyzed using the classification tool based on RDP (98% identity; e-value cutoff of 10⁻³⁰) and M5NR (60% identity; e-value cutoff of 10⁻⁵) with minimum alignment length of 50 bp. The figure displays the taxonomic distribution for the 10 most abundant orders.</p

454 GS FLX Titanium pyrosequencing and Newbler assembly metrics of two metagenomic DNA samples from São Paulo Zoo composting.

<p>454 GS FLX Titanium pyrosequencing and Newbler assembly metrics of two metagenomic DNA samples from São Paulo Zoo composting.</p

ZC1 large contig encoding pectin degradation enzymes.

<p>ZC1 sequences assembled into a 27,919 bp contig encoding the following proteins: 1. Beta-xylosidase (376 aa, COG3507); 2. Dehydrogenases (280 aa, COG1028); 3. hypothetical protein (379 aa); 4. hypothetical protein (283 aa); 5. 5-keto 4-deoxyuronate isomerase (280 aa, COG3717); 6. Dehydrogenases (267 aa, COG1028);7. hypothetical protein (1799 aa); 8. SusD family protein (606 aa, pfam07980); 9. TonB-linked outer membrane protein (1068 aa, COG4771); 10. Pectate lyase (518 aa, COG3866); 11. Predicted unsaturated glucuronyl hydrolase (398 aa, COG4225); 12. Pectin methylesterase (568 aa, COG4677); 13. Endopolygalacturonase (523 aa, COG5434); 14. Nucleoside-diphosphate-sugar epimerase (326 aa, COG0451); 15. Nucleoside-diphosphate-sugar pyrophosphorylase (249 aa, pfam00483); 16. Galactokinase (377 aa, COG0153); 17. Soluble lytic murein transglycosylase (347 aa, COG0741); 18. hypothetical protein (235 aa); 19. Predicted UDP-glucose 6-dehydrogenase (283 aa, COG1004).</p

Features of the composting metagenomes based on MG-RAST^a and IMG/M^b annotations.

a<p>Features from unassembled reads that passed MG-RAST quality control.</p>b<p>Features from Newbler assembled reads post IMG/M data processing.</p><p><b>ND, not determined.</b></p

Top 30 (by sequence count) COG functions represented among ZC1 and ZC2 metagenomic assembled sequences.

<p>793758 and 680461 annotated assembled sequences of ZC1 and ZC2 metagenomes were respectively classified in 4529 and 4872 COGs and ranked according to their abundance. Sequence count and ranking for COGs outside the top 30 list are indicated in numbers with smaller font.</p

Hierarchical clustering of functional gene groups of ZC1 and ZC2 and seven public metagenomes.

<p>(A) Clustering based on COG functional categories; (B) clustering based on COG functions. Hierarchical trees were generated using the “Compare Genomes” tool in IMG/M. Branch lengths are shown.</p

Most abundant bacterial genera in ZC1 and ZC2 compost samples.

<p>Unassembled reads annotated on MG-RAST were analyzed using the classification tool based on RDP (98% identity; e-value cutoff of 10⁻³⁰; minimum alignment length of 50 bp). The figure displays the taxonomic distribution for the 20 most abundant bacterial genera.</p