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Abstract
Inflammatory bowel diseases (IBD) are chronic idiopathic diseases of the gastrointestinal tract and often strike in the prime of life, remaining a lifelong burden. The increasing prevalence of the two main types of IBD, Crohn’s disease (CD) and ulcerative colitis (UC) is indicative of an environmental cause. This thesis explores the mycobacteriology of IBD with emphasis on the gut pathogen, M. avium subspecies paratuberculosis. M. paratuberculosis normally infects livestock and is difficult to detect given its genetic homology with other subspecies of the M. avium complex. In addition, its fastidious growth requirements make isolation from cattle challenging and from sheep and humans extremely difficult, adding to significant controversy as to whether it infects humans. This study demonstrated a significant association between Crohn’s disease and M. paratuberculosis using three PCR assays. Further, the first human M. paratuberculosis ever reported from a patient with UC was isolated. This isolate, 43525, was characterized microbiologically and the genome sequenced on the fourth subculture. M. paratuberculosis 43525 displayed mycobactin dependency on only certain types of media, resistance to; ethambutol, rifampin, clofazamine and streptomycin and growth at 4oC. Given 43525 was a human isolate of what is normally an animal pathogen, the genome sequence provided an opportunity to explore the subtle differences between subspecies of the MAC that infect animals and humans. One PPE along with one PE_PGRS protein was found to be present only in human MAC strains and two PPE/PE genes were unique to 43525 and a caprine M. paratuberculosis strain. The organisation and structure of the 43525 mycobactin cluster differed to other M. paratuberculosis strains providing an explanation for the in vitro phenotype. Finally, a unique mce operon was found to be flanked by two mycobactin genes in all M. paratuberculosis strains investigated. The culture method and validated PCR assays described in this work provide new methods to detect M. paratuberculosis in humans. In addition, the unique mce operon and the PPE/PE proteins highlighted, provide possible targets for diagnostic assays, vaccine candidates or treatment.