Genetics and evolution of Yersinia pseudotuberculosis O-specific polysaccharides: a novel pattern of O-antigen diversity

Abstract

O-antigen polysaccharide is a major immunogenic feature of the lipopolysaccharide of Gram-negative bacteria, and most species produce a large variety of forms that differ substantially from one another. There are 18 known O-antigen forms in the Yersinia pseudotuberculosis complex, which are typical in being composed of multiple copies of a short oligosaccharide called an O unit. The O-antigen gene clusters are located between the hemH and gsk genes, and are atypical as 15 of them are closely related, each having one of five downstream gene modules for alternative main-chain synthesis, and one of seven upstream modules for alternative side-branch sugar synthesis. As a result, many of the genes are in more than one gene cluster. The gene order in each module is such that, in general, the earlier a gene product functions in O-unit synthesis, the closer the gene is to the 5΄ end for side-branch modules or the 3΄ end for main-chain modules. We propose a model whereby natural selection could generate the observed pattern in gene order, a pattern that has also been observed in other species.

Keywords: O antigen; O-specific polysaccharide; Yersinia pseudotuberculosis; gene cluster; lipopolysaccharide; serotype.

Figure 1.

O-antigen gene clusters of Yersinia spp. outside of Y. pseudotuberculosis. Genes are coloured according to the respective pathways of their products, and the scheme is shown on the right. Figure is drawn to scale and the scale is shown below. Bold lines bordering genes indicate that the gene encodes a GT.

Figure 2.

Yersinia pseudotuberculosis O-unit structures. Serotype names are shown above each O unit, and structures are grouped according to common main-chain features. References for structures are in Table 1.

Figure 3.

Yersinia pseudotuberculosis O-antigen gene clusters. Gene clusters are drawn to scale using published sequences (GenBank accession numbers in Table 1). Gene clusters are numerically ordered by serotype name indicated on the left. Genes are coloured according to the respective pathways of their products, and the scheme is shown on the right. Figure is drawn to scale and the scale is shown below. All genes are transcribed from left to right. Bold lines bordering genes indicate that the gene encodes a GT.

Figure 4.

Synthesis pathways of activated sugar precursors that are incorporated into Y. pseudotuberculosis O units. Pathways with experimental data are the CDP-DDH (Chen, Guo and Liu and reviewed in Samuel and Reeves 2003), GDP-l-Colp (Alam, Beyer and Liu 2004), GDP-l-Fucp and GDP-d-Manp (reviewed in Samuel and Reeves 2003) and GDP-6dManHepp (Butty et al. 2009) synthesis pathways. Predicted pathways are for CDP-l-Altf (Cunneen et al. 2009) and GDP-l-Quip (De Castro et al. 2012) synthesis. Substrates and products are shown, and enzymes are in bold face type. Boxed sugars are those incorporated into O units shown in Fig. 1. Sugar abbreviations are expanded in the text.

Figure 5.

Gene modules for O-unit side-branch synthesis in Y. pseudotuberculosis O-antigen gene clusters. Gene cluster details as in legend for Fig. 3. Boxes highlight the gene modules required for the synthesis of side-branch sugars. Names of modules and serotypes are shown on the left. The length of some of these modules can be extended to include the wzx and/or the GT gene, as indicated by the dashed boxes.

Figure 6.

Gene modules for O-unit main-chain synthesis in Y. pseudotuberculosis O-antigen gene clusters. Gene cluster details as in legend for Fig. 3. Boxes highlight the gene modules required for the synthesis of main-chain sugars. Names of modules and serotypes are shown on the left. The length of some of these modules can be extended to include the wzx and/or the GT gene, as indicated by the dashed boxes.

Figure 7.

Phylogenies of Y. pseudotuberculosis wzx ends partitioned at base position 575. Neighbour-joining tree with and bootstrap values based on 100 replicate trees. Serotype names, module groups and wzx names are shown, along with scale bar. Groupings indicated on the right of each phylogeny denote gene immediately upstream or downstream of wzx.

Figure 8.

Pattern representation of the main-chain sugars and their transferases. Each column shows the genes for one of the seven main chains in map order from bottom to top, which puts most of the genes close to function order from top to bottom. The sugars and their linkages are shown in red, below the corresponding GT gene name. GlcpNAc, the first sugar, is shown attached to UndPP at the top, although the wecA gene is not in the gene cluster, followed by rows for the GT genes for the second, third and fourth sugars, which all occur in inverse map order. Intermingled in ascending map order are the other genes in each module. Cells for the initial sugar transferase gene and GT genes are coloured green and cells for other genes are coloured yellow. The significance of the gene order is discussed in the text.

Figure 9.

Comparison of the Y. pseudotuberculosis O:6, O:7 and O:10 gene clusters. Gene cluster details as in legend for Fig. 3. Nucleotide sequence identities are shown between clusters, and the serotype is indicated on the left.