PFGE, Lior serotype, and antimicrobial resistance patterns among Campylobacter jejuni isolated from travelers and US military personnel with acute diarrhea in Thailand, 1998-2003

<p>Abstract</p> <p>Background</p> <p><it>Campylobacter jejuni </it>is a major cause of gastroenteritis worldwide. In Thailand, several strains of <it>C. jejuni </it>have been isolated and identified as major diarrheal pathogens among adult travelers. To study the epidemiology of <it>C. jejuni </it>in adult travelers and U.S. military personnel with acute diarrhea in Thailand from 1998-2003, strains of <it>C. jejuni </it>were isolated and phenotypically identified, serotyped, tested for antimicrobial susceptibility, and characterized using pulsed-field gel electrophoresis (PFGE).</p> <p>Results</p> <p>A total of 312 <it>C. jejuni </it>isolates were obtained from travelers (n = 46) and U.S. military personnel (n = 266) in Thailand who were experiencing acute diarrhea. Nalidixic acid and ciprofloxacin resistance was observed in 94.9% and 93.0% of the isolates, respectively. From 2001-2003, resistance to tetracycline (81.9%), trimethoprim-sulfamethoxazole (57.9%), ampicillin (28.9%), kanamycin (5.9%), sulfisoxazole (3.9%), neomycin (2.0%), and streptomycin (0.7%) was observed. Combined PFGE analysis showed considerable genetic diversity among the <it>C. jejuni </it>isolates; however, four PFGE clusters included isolates from the major Lior serotypes (HL: 36, HL: 11, HL: 5, and HL: 28). The PFGE analysis linked individual <it>C. jejuni </it>clones that were obtained at U.S. military exercises with specific antimicrobial resistance patterns.</p> <p>Conclusions</p> <p>In summary, most human <it>C. jejuni </it>isolates from Thailand were multi-resistant to quinolones and tetracycline. PFGE detected spatial and temporal <it>C. jejuni </it>clonality responsible for the common sources of <it>Campylobacter </it>gastroenteritis.</p

Outer membrane vesicle-mediated release of cytolethal distending toxin (CDT) from Campylobacter jejuni

<p>Abstract</p> <p>Background</p> <p>Background: Cytolethal distending toxin (CDT) is one of the well-characterized virulence factors of <it>Campylobacter jejuni</it>, but it is unknown how CDT becomes surface-exposed or is released from the bacterium to the surrounding environment.</p> <p>Results</p> <p>Our data suggest that CDT is secreted to the bacterial culture supernatant via outer membrane vesicles (OMVs) released from the bacteria. All three subunits (the CdtA, CdtB, and CdtC proteins) were detected by immunogold labeling and electron microscopy of OMVs. Subcellular fractionation of the bacteria indicated that, apart from the majority of CDT detected in the cytoplasmic compartment, appreciable amounts (20-50%) of the cellular pool of CDT proteins were present in the periplasmic compartment. In the bacterial culture supernatant, we found that a majority of the extracellular CDT was tightly associated with the OMVs. Isolated OMVs could exert the cell distending effects typical of CDT on a human intestinal cell line, indicating that CDT is present there in a biologically active form.</p> <p>Conclusion</p> <p>Our results strongly suggest that the release of outer membrane vesicles is functioning as a route of <it>C. jejuni </it>to deliver all the subunits of CDT toxin (CdtA, CdtB, and CdtC) to the surrounding environment, including infected host tissue.</p

Targeted Metabolomics Analysis of Campylobacter coli VC167 Reveals Legionaminic Acid Derivatives as Novel Flagellar Glycans

Glycosylation of Campylobacter flagellin is required for the biogenesis of a functional flagella filament. Recently, we used a targeted metabolomics approach using mass spectrometry and NMR to identify changes in the metabolic profile of wild type and mutants in the flagellar glycosylation locus, characterize novel metabolites, and assign function to genes to define the pseudaminic acid biosynthetic pathway in Campylobacter jejuni 81-176 (McNally, D. J., Hui, J. P., Aubry, A. J., Mui, K. K., Guerry, P., Brisson, J. R., Logan, S. M., and Soo, E. C. (2006) J. Biol. Chem. 281, 18489-18498). In this study, we use a similar approach to further define the glycome and metabolomic complement of nucleotide-activated sugars in Campylobacter coli VC167. Herein we demonstrate that, in addition to CMP-pseudaminic acid, C. coli VC167 also produces two structurally distinct nucleotide-activated nonulosonate sugars that were observed as negative ions at m/z 637 and m/z 651 (CMP-315 and CMP-329). Hydrophilic interaction liquid chromatography-mass spectrometry yielded suitable amounts of the pure sugar nucleotides for NMR spectroscopy using a cold probe. Structural analysis in conjunction with molecular modeling identified the sugar moieties as acetamidino and N-methylacetimidoyl derivatives of legionaminic acid (Leg5Am7Ac and Leg5AmNMe7Ac). Targeted metabolomic analyses of isogenic mutants established a role for the ptmA-F genes and defined two new ptm genes in this locus as legionaminic acid biosynthetic enzymes. This is the first report of legionaminic acid in Campylobacter sp. and the first report of legionaminic acid derivatives as modifications on a protein

Status of vaccine research and development for Campylobacter jejuni

AbstractCampylobacter jejuni is one of the leading causes of bacterial diarrhea worldwide and is associated with a number of sequelae, including Guillain–Barre Syndrome, reactive arthritis, irritable bowel syndrome and growth stunting/malnutrition. Vaccine development against C. jejuni is complicated by its antigenic diversity, a lack of small animal models, and a poor understanding of the bacterium's pathogenesis. Vaccine approaches have been limited to recombinant proteins, none of which have advanced beyond Phase I testing. Genomic analyses have revealed the presence of a polysaccharide capsule on C. jejuni. Given the success of capsule-conjugate vaccines for other mucosal pathogens of global importance, efforts to evaluate this established approach for C. jejuni are also being pursued. A prototypical capsule-conjugate vaccine has demonstrated efficacy against diarrheal disease in non-human primates and is currently in Phase I testing. In addition to proof of concept studies, more data on the global prevalence of capsular types, and a better understanding of the acute and chronic consequences of C. jejuni are needed to inform investments for a globally relevant vaccine

N-Linked Protein Glycosylation Is Required for Full Competence in Campylobacter jejuni 81-176

The recent sequencing of the virulence plasmid of Campylobacter jejuni 81-176 revealed the presence of genes homologous to type IV secretion systems (TFSS) that have subsequently been found in Helicobacter pylori and Wolinella succinogenes. Mutational analyses of some of these genes have implicated their involvement in intestinal epithelial cell invasion and natural competence. In this report, we demonstrate that one of these type IV secretion homologs, Cjp3/VirB10, is a glycoprotein. Treatment with various glycosidases and binding to soybean agglutinin indicated that the structure of the glycan present on VirB10 contains a terminal GalNAc, consistent with previous reports of N-linked glycans in C. jejuni. Site-directed mutagenesis of five putative N-linked glycosylation sites indicated that VirB10 is glycosylated at two sites, N32 and N97. Mutants in the N-linked general protein glycosylation (pgl) system of C. jejuni are significantly reduced in natural transformation, which is likely due, in part, to lack of glycosylation of VirB10. The natural transformation defect in a virB10 mutant can be complemented in trans by using a plasmid expressing wild-type VirB10 or an N32A substitution but not by using a mutant expressing VirB10 with an N97A substitution. Taken together, these results suggest that glycosylation of VirB10 specifically at N97 is required for the function of the TFSS and for full competence in C. jejuni 81-176