Microecology-murine gut-initiation & progression of IBD

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DESCRIPTION (provided by applicant): Genetically engineered mice with immune dysregulation have been used with increasing success in attempting to unravel and comprehend IBD in humans. IBD is considered to be the result of a combination of genetic and environmental factors. Microbial flora are undoubtedly an important component of the disease process, in which microbial antigens are thought to initiate and promote inflammation, particularly in the presence of immune dysregulation or an impaired mucosal barrier in the susceptible host. The understanding of the complex microecology of the distal intestine is extremely limited because of the tedious nature of identifying individual bacterial species and strains by conventional methods, and the inability to culture many fastidious commensal organisms. To circumvent this lack of bacterial speciation, gnotobiotic animals colonized with known microbiota have been used to great advantage. A standardized microbiota used in colonizing germfree rodents referred to "Altered Schaedler Flora" (ASF) was developed. Because of the limitations of an in vivo monitoring system used to identify the 8 anaerobic bacterial species in ASF, we recently characterized their phylogenetic positions relative to known bacteria by utilizing 16S rRNA sequence analysis. This proposal will use molecular techniques, based on quantitative PCR of ASF 1 6S rRNA, to screen the microbial diversity of the murine gut and ascertain how microflora dynamics, under defined experimental conditions, with and without helicobacters, influence initiation and progression of IBD in the mouse. Specifically we will 1) characterize, using molecular techniques, the identification, quantification and distribution of Altered Schaedler's Flora (ASF) in the gastrointestinal tract of the IL-1O-/- and the C57BL mouse. 2) Determine how enteropathogenic Helicobacter spp. associated typhlocolitis in the lL-1O-/- mice alters the microecology of the lower bowel and correspondingly, ascertain whether and how individual species in ASF influence the progression or attenuation of chronic intestinal inflammation. 3) Determine microbial dynamics of ASF after physiological and anatomical perturbation of the lower bowel in mice as well as altering host genotype an how these changes influence subsequent induction of typhlocolitis and possibly colonic adenocarcinoma induced by Helicobacter spp. 4) Determine perturbations of ASF and Helicobacter spp. prior and subsequent to oral vaccination with Helicobacter spp. antigens and mucosal adjuvants, elucidate how these vaccine strategies influence microflora dynamics and impact gut cytokine responses.