Identification of putative virulence factors from Campylobacter spp. isolated in Iceland

• Main Author: Akins, Edith Deann
• Format: Doctoral or Postdoctoral Thesis
• Language: English
• Published: uga 2008
• Subjects: Campylobacter; virulence; capacitance; Bactometer; cell invasion; microarray hybridization; suppressive subtractive hybridization; Kelli; Mueller; Iceland
• Online Access: http://hdl.handle.net/10724/24812; http://purl.galileo.usg.edu/uga_etd/akins_edith_d_200808_phd

Two studies were conducted to determine putative virulence factors of Campylobacter spp. that were isolated in Iceland. The first study investigated capacitance monitoring using a simplified medium for efficient and reproducible construction of growth curves for Campylobacter spp., which can be a time consuming and labor intensive process. When invasion assays are performed, it is required that Campylobacter spp. isolates be grown to a 68density of 10 to 10 CFU/ml. This investigation optimized conditions for use with the Bactometer® such that the determination of growth curves was achieved in a simple medium. Results suggested that isolates should be grown on Mueller Hinton plates under a microaerobic atmosphere (37¡C; 24 h), then transferred to Mueller Hinton biphasic cultures for 6 h (37¡C; microaerobic atmosphere). Serial dilutions should be used for inoculation of Bactometer® wells containing 1 mL Mueller Hinton broth plus 0.1M sodium pyruvate for obtaining growth curves. In the second study, putative virulence factors of Campylobacter spp. were investigated. Campylobacter spp. exhibited a wide distribution of adhesion and invasion ability, which was determined to be unrelated to flaA short variable region allele type. The second part of this study investigated the most invasive isolate 14118, the least invasive isolate, 13262, and two in between to further understanding of the molecular basis of genetic diversity among these 4 C. jejuni isolates. DNA-DNA microarray hybridizations identified genes absent relative to C. jejuni 11168 (PMSRU). Several absent genes were located in 1 of 7 previously described plasticity regions. There were 372 genes determined to be present in C. jejuni isolates 14118, 5116, 8557 and 13262 as well as C. jejuni 11168 (PMSRU). DNA suppressive subtractive hybridizations identified genes not in common with C. jejuni 11168 (PMSRU). C. jejuni 14118 contained a gene from C. doylei 269.97 that encoded for a motility accessory factor. C. jejuni 13262 contained a cytolethal distending toxin operon from C. lari. as well as a type II restriction modification enzyme unlike isolate 14118, 5116 and 8557 which includes a type I restriction modification enzyme. PhD Food Science and Technology Food Science Mark Harrison Mark Harrison Jinru Chen Mark Berrang Joseph Frank Kelli Hiett