Characterization of the Burkholderia pseudomallei K96243 Capsular Polysaccharide I Coding Region

Abstract

Burkholderia pseudomallei is the causative agent of melioidosis, a disease endemic to regions of Southeast Asia and Northern Australia. Both humans and a range of other animal species are susceptible to melioidosis, and the production of a group 3 polysaccharide capsule in B. pseudomallei is essential for virulence. B. pseudomallei capsular polysaccharide (CPS) I comprises unbranched manno-heptopyranose residues and is encoded by a 34.5-kb locus on chromosome 1. Despite the importance of this locus, the role of all of the genes within this region is unclear. We inactivated 18 of these genes and analyzed their phenotype using Western blotting and immunofluorescence staining. Furthermore, by combining this approach with bioinformatic analysis, we were able to develop a model for CPS I biosynthesis and export. We report that inactivating gmhA, wcbJ, and wcbN in B. pseudomallei K96243 retains the immunogenic integrity of the polysaccharide despite causing attenuation in the BALB/c murine infection model. Mice immunized with the B. pseudomallei K96243 mutants lacking a functional copy of either gmhA or wcbJ were afforded significant levels of protection against a wild-type B. pseudomallei K96243 challenge.

Neutrophil Extracellular Traps Exhibit Antibacterial Activity against Burkholderia pseudomallei and Are Influenced by Bacterial and Host Factors

“…The response was greater with a wcbB mutant completely lacking the polymer of unbranched manno-heptopyranose residues than with a wcbN mutant that retains a capsule-specific epitope but whose virulence is attenuated (9). In common with other encapsulated bacterial pathogens, it appears that the B. pseu-…”

“…4A). In addition, we observed that the amount of extracellular DNA released by human PMNs was significantly higher following infection by a wcbB mutant which lacks capsular polysaccharide I (CPS-I; a chromosome 1-encoded polymer of unbranched manno-heptopyranose residues [9]) than infection by the K96243 parent strain (Fig. 4A).…”

“…4A). Upon infection by a wcbN mutant, which lacks the ability to synthesize GDP-Dglycero-␣-D-manno-heptopyranose and retains an epitope for a capsule-specific antibody but has reduced virulence in mice (8,9), the amount of extracellular DNA released by PMNs was elevated relative to that released by cells infected by the parent strain, but not significantly so (Fig. 4A).…”

Determination of Protein-ligand Interactions Using Differential Scanning Fluorimetry

“…The putative heptose-guanyl transferase WcbM 19,20 shows a strong thermal shift on binding to GTP ( Figure 3A). An initial screen suggested that the K d would be in the range of around 100 µM.…”

Reliability of Automated Biochemical Identification of Burkholderia pseudomallei Is Regionally Dependent

“…N-Acetylgalactosamine, a derivative of NAGA, has also been documented as one of the basic components for EPS of B. pseudomallei (19). The implications for virulence and immune response of these different biochemical profiles remains uncertain, but it has been suggested that the amount of capsular polysaccharide in B. pseudomallei compared to that in other Burkholderia species may well contribute to its relative virulence (20).…”