The type II O‐antigenic polysaccharide moiety of Burkholderia pseudomallei lipopolysaccharide is required for serum resistance and virulence

Abstract

SummaryMelioidosis, an infection caused by the Gram-negative bacterial pathogen Burkholderia pseudomallei, is endemic in south-east Asia and northern Australia. Acute septicaemic melioidosis is a major cause of morbidity and mortality, especially in north-east Thailand. B. pseudomallei is highly resistant to the bactericidal activity of normal human serum (NHS), and we have found that B. pseudomallei 1026b multiplies in 10-30% NHS. We developed a simple screen for the identification of serum-sensitive mutants based on this novel phenotype. Approximately 1200 Tn5-OT182 mutants were screened, and three serum-sensitive mutants were identified. The type II O-antigenic polysaccharide (O-PS) moiety of lipopolysaccharide was not present in the serum-sensitive mutants. A representative serum-sensitive mutant, SRM117, was killed by the alternative pathway of complement and was less virulent than 1026b in three animal models of melioidosis. The Tn5-OT182 integrations in the serum-sensitive mutants were physically linked on the B. pseudomallei chromosome, and further genetic analysis of this locus revealed a cluster of 15 genes required for type II O-PS production. The proteins encoded by these genes were similar to proteins involved in bacterial polysaccharide biosynthesis.The results presented here demonstrate that type II O-PS is essential for B. pseudomallei serum resistance and virulence.

Characterisation of an acapsular mutant of Burkholderia pseudomallei identified by signature tagged mutagenesis

“…B. pseudomallei produces two types of lipopolysaccharide (LPS) termed OPSI and OPSII and a capsular polysaccharide [20,21]. OPSII appears to play a role in resistance to serum killing and virulence in the diabetic rat model of infection [22,23]. The polysaccharide capsule also appears to be an important virulence factor, and a transposon mutant that failed to produce exopolysaccharide was markedly attenuated in Syrian hamsters [24].…”

Cloning and analysis of genomic differences unique to Burkholderia pseudomallei by comparison with B. thailandensis

“…Of these, only protease has been shown to be involved in pathogenesis, with a protease-de®cient mutant having a moderately increased LD50 in an animal model [4]. The type II Oantigenic polysaccharide moiety of lipopolysaccharide is a demonstrated virulence factor [6], whereas a number of incompletely characterised toxins, and a type III secretion apparatus, show potential as virulence factors [7±9].…”

Identification of an ATP-binding Cassette Transporter for Export of the O-antigen across the Inner Membrane inRhizobium etli Based on the Genetic, Functional, and Structural Analysis of an lps Mutant Deficient in O-antigen

“…The predicted gene product is similar to the KpsM-like protein of Synechocystis sp. (27% amino acid identity), Wzm of P. aeruginosa (22% identity), and Wzm of B. pseudomallei (22% identity) (43). As for Wzt, we postulate the existence of two classes of Wzm-like proteins based on the results of a phylogenetic analysis (Fig.…”

“…Therefore, the isolation of a wzt-homologous gene responsible for the export of such a heteropolymeric Oantigen in R. etli CE3 is of interest since most of the bacterial polysaccharide biosynthetic genes encoding ABC-2 transporters are involved in the biosynthesis of homopolysaccharides (6). Other known exceptions are the gene cluster of A. actinomycetemcomitans involved in the biosynthesis of the heteropolymeric serotype b-specific polysaccharide antigen and the heteropolymeric type II O-polysaccharide gene cluster of B. pseudomallei, which also contains the wzm and wzt genes (43,52).…”