Content area
Abstract
Campylobacter is a zoonotic bacterium and is the main cause of bacterial foodborne gastroenteritis worldwide, with most human campylobacteriosis cases being the result of infection with Campylobacter jejuni or Campylobacter coli.Transmission via handling and consumption of (poultry) meat only contributes to half of all campylobacteriosis cases, highlighting other transmission routes such as contact with live animals, contact with pets, and via environmental waters.
C. jejuni and C. coli were previously considered asaccharolytic, but are now known to carry specific saccharides metabolization pathways, the so-called L-fucose and Dglucose utilization clusters. Approximately 52% and 75% of the investigated C. jejuni and C. coli isolates carry the L-fucose utilization cluster (Cj0480c – Cj0489), while 0.25% and 6.3% carry the D-glucose utilization cluster (glcP, pgi2 glk, pgl, zwf, edd and eda), respectively. This thesis describes research on Campylobacter to investigate whether these clusters are beneficial for Campylobacterniche adaptation and help establish human infection.
In Chapter 2 the L-fucose utilization cluster in C. jejuni and C. coli isolates is characterized including roles in growth, survival and metabolism. After incubation for seven days, comparative analysis of performance in L-fucose-enriched MEMα medium compared to MEMα, showed an enhanced survival and prolonged spiral cell morphology for all tested isolates, with the C. coli isolate having the most robust phenotype. Furthermore, HPLC analysis indicated that L-fucose utilization was linked to acetate, lactate, pyruvate and succinate production. Genetic analysis of the L-fucose utilization cluster revealed a frameshift in the Cj0489 gene, resulting in two open reading frames encoding for putative 77 amino acid N-terminal (Cj0489-S) and 394 amino acid C-terminal (Cj0489-L) proteins, in the reference isolate C. jejuniNCTC11168, which did not result in a different phenotype of growth, survival and metabolism compared to the other tested Campylobacter isolates.
Chapter 3 describes an extended phenotypic testing of the L-fucose utilization cluster in two closely related human C. jejuni isolates, C. jejuni NCTC11168 and C. jejuni 286, and next to L-fucose metabolism, also D-arabinose metabolism was studied. Darabinose is a sugar that is similar to L-fucose and was found to be metabolized via the L-fucose utilization cluster of C. jejuni. L-fucose and D-arabinose metabolism, including metabolite production, and its impact on Caco-2 cell interaction and binding to fibronectin were investigated. Increased survival and changes in metabolism were observed in both tested isolates when cultured with L-fucose and Darabinose, and resulted in production of acetate, pyruvate and succinate, and the respective signature metabolites lactate and glycolic acid, which was in line with the upregulation of L-fucose cluster proteins. In vitro Caco-2 cell studies and fibronectinbinding experiments showed significantly higher invasion and fibronectin binding efficacy of C. jejuni NCTC11168 cells grown with L-fucose or D-arabinose, compared to cells grown without these substrates, while no significant differences were found with C. jejuni286 between all tested media. The fibronectin binding proteins CadF and FlpA were both detected in the two isolates, however, not significantly differentially expressed in cells grown with L-fucose or D-arabinose.
