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Reconstitution of O-Specific Lipopolysaccharide Expression in Burkholderia cenocepacia Strain J2315, Which Is Associated with Transmissible Infections in Patients with Cystic Fibrosis

Ximena Ortega,¹ Tracey A. Hunt,^1, Slade Loutet,¹ Arlene D. Vinion-Dubiel,² Anup Datta,³ Biswa Choudhury,³ Joanna B. Goldberg,² Russell Carlson,^3,4 and Miguel A. Valvano^1*

Department of Microbiology and Immunology, University of Western Ontario, London, Ontario, Canada,¹ Department of Microbiology, University of Virginia Health System, Charlottesville, Virginia,² Department of Biochemistry and Molecular Biology,³ Complex Carbohydrate Research Center, University of Georgia, Athens, Georgia⁴

Received 9 August 2004/ Accepted 17 November 2004

Burkholderia cenocepacia is an opportunistic bacterium that infects patients with cystic fibrosis. B. cenocepacia strains J2315, K56-2, C5424, and BC7 belong to the ET12 epidemic clone, which is transmissible among patients. We have previously shown that transposon mutants with insertions within the O antigen cluster of strain K56-2 are attenuated for survival in a rat model of lung infection. From the genomic DNA sequence of the O antigen-deficient strain J2315, we have identified an O antigen lipopolysaccharide (LPS) biosynthesis gene cluster that has an IS402 interrupting a predicted glycosyltransferase gene. A comparison with the other clonal isolates revealed that only strain K56-2, which produced O antigen and displayed serum resistance, lacked the insertion element inserted within the putative glycosyltransferase gene. We cloned the uninterrupted gene and additional flanking sequences from K56-2 and conjugated this plasmid into strains J2315, C5424, and BC7. All the exconjugants recovered the ability to form LPS O antigen. We also determined that the structure of the strain K56-2 O antigen repeat, which was absent from the LPS of strain J2315, consisted of a trisaccharide unit made of rhamnose and two N-acetylgalactosamine residues. The complexity of the gene organization of the K56-2 O antigen cluster was also investigated by reverse transcription-PCR, revealing several transcriptional units, one of which also contains genes involved in lipid A-core oligosaccharide biosynthesis.

Supplemental material for this article may be found at http://jb.asm.org/.

Present address: Department of Cell Biology, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada.

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