Characterization of theCampylobacter jejuniHeptosyltransferase II Gene,waaF, Provides Genetic Evidence that Extracellular Polysaccharide Is Lipid A Core Independent

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Abstract
Campylobacter jejuniproduces both lipooligosaccharide (LOS) and a higher-molecular-weight polysaccharide that is believed to form a capsule. The role of these surface polysaccharides inC. jejuni-mediated enteric disease is unclear; however, epitopes associated with the LOS are linked to the development of neurological complications. InEscherichia coliandSalmonella entericaserovar Typhimurium thewaaFgene encodes a heptosyltransferase, which catalyzes the transfer of the secondl-glycero-d-manno-heptose residue to the core oligosaccharide moiety of lipopolysaccharide (LPS), and mutation ofwaaFresults in a truncated core oligosaccharide. In this report we confirm experimentally thatC. jejunigene Cj1148 encodes the heptosyltransferase II enzyme, WaaF. TheCampylobacter waaFgene complements anS. entericaserovar TyphimuriumwaaFmutation and restores the ability to produce full-sized lipopolysaccharide. To examine the role of WaaF inC. jejuni,waaFmutants were constructed in strains NCTC 11168 and NCTC 11828. Loss of heptosyltransferase activity resulted in the production of a truncated core oligosaccharide, failure to bind specific ligands, and loss of serum reactive GM1, asialo-GM1, and GM2ganglioside epitopes. The mutation ofwaaFdid not affect the higher-molecular-weight polysaccharide supporting the production of a LOS-independent capsular polysaccharide byC. jejuni. The exact structural basis for the truncation of the core oligosaccharide was verified by comparative chemical analysis. The NCTC 11168 core oligosaccharide differs from that known for HS:2 strain CCUG 10936 in possessing an extra terminal disaccharide of galactose-β(1,3)N-acetylgalactosamine. In comparison, thewaaFmutant possessed a truncated molecule consistent with that observed withwaaFmutants in other bacterial species.