The role of Campylobacter concisus in enteric infections
Creators: name: Elshagmani, Eltahar
Publisher: name: RMIT University
Year: 2024

description: Campylobacter concisus is a fastidious hydrogen-requiring, Gram-negative, curved rod. This bacterium can be frequently isolated from the oral cavity of healthy individuals; however, the first isolation of C. concisus was in 1981 from a gum infection and it was then suggested to be a potential cause of gastroenteritis in children and immunocompromised adults. More recently, C. concisus has been associated with inflammatory bowel diseases (IBD) in children. However, some studies could not report a significant difference in the prevalence of C. concisus between IBD patients and healthy controls.<br><br>The potential pathogenic role of C. concisus in enteric infections is still not well investigated. Additionally, it has not been confirmed yet whether C. concisus is an opportunistic pathogen or if a certain group/genomospecies can cause human infections. Moreover, the virulence of C. concisus isolates from the oral cavity of healthy individuals and intestinal isolates has not been intensely investigated.<br><br>C. concisus has been grouped to two different genomospecies (A and B) on the basis of DNA-DNA hybridisation studies and PCR amplification of 23S rDNA. This study investigated the molecular diversity of potential pathogenic groups of C. concisus using two molecular techniques: PCR-denaturing gradient gel electrophoresis (PCR-DGGE) (to amplify and analyse 16S rDNA), and a conventional PCR (to amplify several putative virulence genes including cjaC, cjaA, dnaJ and zot). These techniques were applied to C. concisus faecal and oral isolates. DGGE grouped C. concisus isolates into two new distinct groups. The putative virulence genes, cjaC, cjaA and dnaJ, were detected in the majority of C. concisus isolates, but there were DNA sequence variations, which could have led to PCR amplification failure in some C. concisus isolates. It was found that the location of cjaC in the genome of genomospecies A was different from its location in the genome of genomospecies B. Another virulence gene (zot) encodes a tight junction toxin, could be only amplified from a few isolates.<br><br>An invasion assay was used to investigate the virulence of C. concisus using the human intestinal cell line INT407. Two C. concisus isolates from the oral cavity of healthy volunteers were identified as invasive isolates, while all tested faecal isolates from patients with diarrhoea were found to be non-invasive. Oral isolates obtained from healthy volunteers have not been previously reported to be more invasive than clinical intestinal isolates. Furthermore, C. concisus isolates possessing zot were unable to show significant invasion capability. In addition, semi-quantitative SYBR green assays were developed to evaluate the expression of the putative virulence genes (cjaC, cjaA, dnaJ and zot) in C. concisus grown on Columbia agar with the addition of horse blood (HBA), on Columbia agar (CA) only, maintained in tissue culture medium with or without INT407 cells. The expression of cjaC and cjaA was slightly enhanced when the bacterial cells were grown on CA compared with HBA. However, dnaJ expression was significantly reduced in bacterial cells sustained in the tissue culture medium only when compared with those maintained with INT407 cells growing in a similar tissue culture medium.<br><br>Another aspect of this study was to investigate the prevalence and possible association of particular C. concisus genomospecies in adults with IBD. A total of 350 intestinal biopsies (at least seven intestinal biopsies from each participant) were collected from 51 participants (IBD and control participants) attending the Gastroenterology Clinic at the Austin Hospital in Melbourne, Australia. Also from each participant, a gum and faecal samples were collected at the time of the biopsy collection. The presence of C. concisus was investigated by conventional culture and molecular techniques in all collected specimens. To detect the prevalence of C. concisus genomospecies B and zot directly from the clinical samples, new nested PCR techniques were developed. The direct detection of genomospecies A DNA in the clinical samples was not included, as in the start of this study the whole genome of genomospecies A has not been published.<br><br>Overall, the prevalence of C. concisus DNA was significantly higher in biopsies of IBD patients (59.5%) than in biopsies of control participants (21.4%). Moreover, C. concisus was isolated for the first time from duodenal and ascending colonic biopsies. The prevalence of genomospecies B was significantly higher in IBD patients than in control participants using the nested PCR to detect this genomospecies in intestinal biopsies, faeces and gum samples. Among the C. concisus isolates cultured from gum samples, the prevalence of genomospecies B was also significantly higher in IBD patients than in control participants, while genomospecies A was more predominant in control participants. These findings suggested that genomospecies B might be associated with IBD. Furthermore, zot was not detected in most of the clinical isolates and there was no significant difference in its prevalence between IBD patients and control participants.<br><br>Collectively, the main results of this research project reinforced the extreme genetic diversity of C. concisus and more specifically between genomospecies A and B, on the basis of the putative virulence genes. In addition, it was shown that C. concisus invasive strains can exist in the oral cavity of healthy individuals. Gene expression assays for cjaC, cjaA and dnaJ were successfully developed in this study for the first time and it was shown that dnaJ was possibly involved in cellular invasion since its expression significantly reduced after exposure to tissue culture medium without INT407 cells. Furthermore, new PCR techniques were developed to detect genomospecies B and zot directly from clinical samples. The results suggested an association of C. concisus, in particular genomospecies B, with adults with IBD while no association of zot was recognised in the same group.