Characterization of two Campylobacter jejuni strains for use in volunteer experimental-infection studies

Abstract

The development of vaccines against Campylobacter jejuni would be facilitated by the ability to perform phase II challenge studies. However, molecular mimicry of the lipooligosaccharide (LOS) of most C. jejuni strains with human gangliosides presents safety concerns about the development of Guillain-Barré syndrome. Clinical isolates of C. jejuni that appeared to lack genes for the synthesis of ganglioside mimics were identified by DNA probe analyses. Two clinical isolates from Southeast Asia (strains BH-01-0142 and CG8421) were determined to express the LOS type containing N-acetyl quinovosamine. No ganglioside structures were observed to be present in the LOSs of these strains, and pyrosequence analyses of the genomes of both strains confirmed the absence of genes involved in ganglioside mimicry. The capsule polysaccharide (CPS) of BH-01-0142 was determined to be composed of galactose (Gal), 6-deoxy-ido-heptose, and, in smaller amounts, D-glycero-D-ido-heptose, and the CPS of CG8421 was observed to contain Gal, 6-deoxy-altro-heptose, N-acetyl-glucosamine, and minor amounts of 6-deoxy-3-O-Me-altro-heptose. Both CPSs were shown to carry O-methyl-phosphoramidate. The two genomes contained strain-specific zones, some of which could be traced to a plasmid origin, and both contained a large chromosomal insertion related to the CJEI3 element of C. jejuni RM1221. The genomes of both strains shared a high degree of similarity to each other and, with the exception of the capsule locus of CG8421, to the type strain of the HS3 serotype, TGH9011.

Figures

FIG. 1.

LOS structure and genetic loci. (A) LOS structures of CG8421 and BH-01-0142. The asterisks indicate moieties that were present in nonstoichiometric amounts. (B) LOS locus of CG8421. ORFs are labeled by both their gene names and the nomenclature of Parker et al. (43), in which “h” designates the H LOS class, exemplified by TGH9011, and “e” designates the E LOS class, exemplified by 81116. Genes marked with an asterisk contain homopolymeric tracts of G or C. The LOS locus of BH-01-0142 is identical to that of CG8421, with two exceptions. Orf28e (in gray) is missing in BH-01-0142, and Orf31h of CG8421, but not that of BH-01-0142, contains a frameshift, as discussed in the text.

FIG. 2.

1H NMR spectrum of C. jejuni BH-01-0142 core OS showing the deoxy resonances of QuiNAc and the characteristic N-acetyl resonances of QuiNAc, GlcNAc, and GalNAc. HOD, water.

FIG. 3.

31P NMR spectrum of C. jejuni CG8421 core OS showing the presence of two monoester phosphate moieties.

FIG. 4.

GC-MS profile of the alditol acetate derivatives of BH-01-0142 CPS (A) and CG8421 CPS (B) showing the monosaccharide residues present in their respective CPSs.

FIG. 5.

Comparison of the DNA of the capsule loci of C. jejuni strains 81-176 and CG8421. A comparison was made using the Artemis comparison tool software (http://www.sanger.ac.uk/Software/ACT/). The vertical block between CPS sequences represents conservation. MeOPN, methyl phosphoramidate.

FIG. 6.

Comparison of the DNA of the flagellin glycosylation loci of C. jejuni strains BH-01-0142, CG8421, and NCTC 11168. A comparison was made using Artemis comparison tool software (http://www.sanger.ac.uk/Software/ACT/). The vertical block between sequences represents conservation. Flagellin glycosylation sequences from BH-01-0142 and CG8421 are incomplete and are represented by 15 and 7 contigs, respectively. The order of the contigs presented is based on the alignment with the NCTC 11168 locus and might not reflect the physical order in BH-01-0142 and CG8421.