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. 2010 Jan;192(2):525-38.
doi: 10.1128/JB.01144-09. Epub 2009 Nov 6.

The Citrobacter Rodentium Genome Sequence Reveals Convergent Evolution With Human Pathogenic Escherichia Coli

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The Citrobacter Rodentium Genome Sequence Reveals Convergent Evolution With Human Pathogenic Escherichia Coli

Nicola K Petty et al. J Bacteriol. .
Free PMC article

Abstract

Citrobacter rodentium (formally Citrobacter freundii biotype 4280) is a highly infectious pathogen that causes colitis and transmissible colonic hyperplasia in mice. In common with enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively), C. rodentium exploits a type III secretion system (T3SS) to induce attaching and effacing (A/E) lesions that are essential for virulence. Here, we report the fully annotated genome sequence of the 5.3-Mb chromosome and four plasmids harbored by C. rodentium strain ICC168. The genome sequence revealed key information about the phylogeny of C. rodentium and identified 1,585 C. rodentium-specific (without orthologues in EPEC or EHEC) coding sequences, 10 prophage-like regions, and 17 genomic islands, including the locus for enterocyte effacement (LEE) region, which encodes a T3SS and effector proteins. Among the 29 T3SS effectors found in C. rodentium are all 22 of the core effectors of EPEC strain E2348/69. In addition, we identified a novel C. rodentium effector, named EspS. C. rodentium harbors two type VI secretion systems (T6SS) (CTS1 and CTS2), while EHEC contains only one T6SS (EHS). Our analysis suggests that C. rodentium and EPEC/EHEC have converged on a common host infection strategy through access to a common pool of mobile DNA and that C. rodentium has lost gene functions associated with a previous pathogenic niche.

Figures

FIG. 1.
FIG. 1.
Phylogeny of C. rodentium showing the phylogenetic relationship of C. rodentium to various enteric bacteria based on the nucleotide sequences of seven housekeeping genes (adk, fumC, gyrB, icd, mdh, purA, and recA). The tree shows bootstrap values (percentages of 1,000 replicates) below the branches and was rooted using an outgroup comprising Yersinia, Serratia, and Pectobacterium. The posterior probability for each node was 1 in every case and thus is not shown on the tree. The scale bar represents the number of substitutions per site.
FIG. 2.
FIG. 2.
Orthologous CDSs in C. rodentium ICC168, E. coli E2348/69 (EPEC), and E. coli O157:H7 Sakai (EHEC). The Venn diagram shows the number of genes that are unique to one strain, or shared between two or three of the strains, based on the results of reciprocal FASTA analysis with a minimum similarity of 40% identity over 80% of the CDSs. The numbers in parentheses indicate the numbers of C. rodentium genes in that category that have no orthologue in E. coli K-12 strain MG1655.
FIG. 3.
FIG. 3.
Circular map of the C. rodentium ICC168 chromosome. From the outside in, the first circle shows the positions of genomic islands and prophages (detailed in Table 3). The second circle shows the genomic positions in Mbp. The third and fourth circles show the CDSs transcribed clockwise and counterclockwise, respectively (color coded according to the predicted functions of their gene products: black, pathogenicity or adaptation; gray, energy metabolism; red, information transfer; green, membrane or surface structure; yellow, central or intermediary metabolism; cyan, degradation of macromolecules; cerise, degradation of small molecules; pale blue, regulator; pink, prophage or IS element; orange, conserved hypothetical; pale green, unknown; and brown, pseudogene). The fifth circle shows C. rodentium CDSs (dark blue) that lack orthologues (by reciprocal FASTA analysis) in EPEC E2348/69, EHEC Sakai, or K-12 MG1655. The sixth circle shows C. rodentium CDSs (black) that have orthologues (by reciprocal FASTA analysis) in both EPEC and EHEC but not K-12. Circles 7, 8, and 9 show the positions of C. rodentium CDSs that have orthologues (by reciprocal FASTA analysis) in EPEC (red), EHEC (green), or K-12 (orange) (excluding those CDSs that also had orthologues in one or both of the other E. coli strains). The innermost circle shows a plot of G+C content.
FIG. 4.
FIG. 4.
EspS is a T3SS translocated protein. Translocation of EspS-TEM-1 and the control EspH-TEM-1 from wild-type (ICC169) and ΔespA (ICC304) C. rodentium was quantified using a Fluostar Optima reader with excitation at 410 nm (10-nm band-pass). Emission was detected via 450-nm (blue fluorescence) and 520-nm (green fluorescence) filters. The translocation rate was expressed as the 450/520-nm emission ratio. EspS and EspH were translocated from wild-type C. rodentium, but not from the ΔespA mutant. The error bars represent mean standard deviations (SD).
FIG. 5.
FIG. 5.
Colonization dynamic of wild-type, ΔespA (ICC304), and ΔespA(pACYC-espA) C. rodentium strains after oral inoculation of C3H/Hej mice. Mice inoculated with wild-type (WT) and ΔespA(pACYC-espA) C. rodentium exhibited similar colonization dynamics, whereas ΔespA C. rodentium was unable to colonize. The inoculum count is the number of viable bacteria in 200 ml used to inoculate mice by oral gavage. The data are represented as means ± SD.
FIG. 6.
FIG. 6.
T6SSs in C. rodentium and EHEC. Shown is a schematic representation of the C. rodentium ICC168 T6SS gene clusters CTS1 and CTS2 and the EHEC O157:H7 Sakai T6SS cluster EHS. Conserved T6SS components are highlighted. The differing architectures and lack of homology between CTS1, CTS2, and EHS suggest that these T6SS clusters have distinct evolutionary histories.

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