Advanced Metagenomic Analysis of Human Colonic Microbiota in Patients With Chronic GI Disorders
Condition: Irritable Bowel Syndrome; Colitis, Ulcerative; Crohn Disease; Colonic Neoplasms
MeSH terms: Colonic Neoplasms; Crohn Disease; Irritable Bowel Syndrome; Colitis, Ulcerative
Study type: Observational

Summary: 
      This clinical trial hypothesize that Gut Microbiota (bacteria, viruses, fungi)play a major
      role in the occurrence and progression of many chronic gastrointestinal diseases like
      Irritable Bowel Syndrome, Inflammatory Bowel Diseases and Colo-Rectal Cancer.

      Hence, aims to study the spectrum of such microbiota in these patients as compared to normal
      subjects, by utilizing metagenomic techniques rather than cultural methods.
    

Detailed description: 
      The terms intestinal "microflora" or "microbiota" refer to the microbial ecosystem colonizing
      the gastrointestinal tract. Recently developed molecular biology instruments suggest that a
      substantial part of bacterial communities within the human gut still have to be described.
      Intestinal bacteria play an essential role in the development and homeostasis of the immune
      system. Most of these important microbiota are unculturable which significantly have limited
      our understanding of bacterial-host crosstalk. Among the methods designed to gain access to
      the physiology and genetics of uncultured organisms, metagenomics, the genomic analysis of a
      population of microorganisms, has emerged as a powerful technology. Metagenomics is the study
      of genomic content in a complex mixture of microorganisms. Direct isolation of genomic DNA
      from an environment circumvents culturing the organisms under study. The two primary goals of
      this approach are to develop a consensus of what populations of microorganisms are present
      and then to identify what roles each microorganism has within a specific environment.
      Metagenomics samples are found nearly everywhere, including several microenvironments within
      the human gut, soil samples, extreme environments such as deep mines and the various layers
      within the ocean. Therefore, the diversity of microorganisms is thought to be in the range of
      hundreds of millions to greater than tens of trillions of species. Among the most mysterious
      microenvironment is the human gastrointestinal tract that harbor greater than thousands of
      millions of microbial species (at least 1014), including up to 2000 species dominated by
      anaerobic bacteria.

      Many of the gastrointestinal or even other diseases (metabolic as in obesity, or autoimmune
      as in allergies) involve primarily the human gut microbiota and then according to specific
      changes in microbiota equilibrium certain effects occur on either bowel motility (as in
      irritable bowel syndrome: IBS), homeostasis of the GI immune system (as in inflammatory bowel
      disease: IBD), or mucosal cells proliferation (as in adenoma - colorectal cancer: CRC). These
      chronic diseases affect all nations worldwide and represent a significant public health
      burden. They can be seen among children, adolescents, and adults. Currently there is no
      medical cure for IBS, IBD or CRC once they develop.

      The complex interactions between microbial, genetic, immune, and environmental factors seem
      to play an important role in the pathogenesis of IBS, IBD and CRC. Lately, post infectious -
      IBS have gained increasing focus, to the extent that whole pathogenesis of IBS might be
      attributed to a specific triggering factor of microbiota imbalance. The prevailing theory is
      that IBD is related to an altered mucosal barrier with a deregulated immune response directed
      against specific modifications in the normal microbiota leading to the alteration of its
      equilibrium. The etiology of IBD can therefore be conceptualized as an aberrant immunologic
      response to a modified component or components of the gut microbiota potentially following an
      environmental insult. Likewise, CRC development process from normal mucosal surface to
      adenoma and finally to CRC; is probably related to gut microbiota.

      The prevalence of these diseases has been documented to go through an obvious increase in
      Saudi Arabia during the last 3 decades. This would represent a unique model to study the role
      of GIT microbiota or their metagenomics and their modifications in response to environmental
      or dietary factors in this community that went into urbanization fairly recently, and then
      analyze their causative relations to the focus diseases.

      Here, we propose to perform a comprehensive analysis of the gastrointestinal tract microbiota
      and its contribution on gut homeostasis in normal subjects and patients with IBS, IBD and CRC
      by using state of the art metagenomics technology. This will be done on a Saudi population
      sample that we believe represent a unique model.

      Our specific objectives for this project are:

        -  Characterize the microbiota composition (microbes and virus) of the mucosa from Saudi
           patients with IBS, IBD and CRC.

        -  Characterize the mobile GI metagenomics of Saudi patients with IBS, IBD and CRC.

        -  Compare the metagenomics of IBS, IBD and CRC patients to each other and to normal
           subjects from the same population.

      Expected outcomes and Significance of research:

      Altogether, the results from aims 1 and 2 will provide for the first time a comprehensive and
      in-depth analysis of the mucosa-associated microbiota of adult patients with IBS, IBD and CRC
      in Saudi population. The proposed study will define a microbiota "fingerprint" for Saudi
      norms, IBS, IBD and CRC. The contribution of virome and the mobile metagenome into these
      diseases development and/or health maintenance will be assessed for the first time and thus
      has the potential to reveal new paradigms. In addition, the study of the virome and mobile
      metagenome will help us to understand the selective forces that could contribute to the
      alteration and evolution of the microbiota community and thus could have important
      implications for the treatment of the diseases. Certainly, the work proposed here will pave
      the way toward future hypothesis-driven research which could lead to the design of
      therapeutic strategies aimed at manipulating the microbial community.
    

Observational model: Case-Control
Time perspective: Cross-Sectional
Sampling method: Non-Probability Sample
Study population: 
        We propose to recruit our cohort of patients from the King Fahad Medical City in Riyadh
        (largest tertiary care hospital of Saudi MOH with wide open referral system), over one year
        between July 2010 and July of 2011. Fifty participants with each of the focus conditions
        (IBS, UC, CD, CRC) will be compared to fifty participants with no GI complaints and have a
        normal colonic mucosa (by endoscopic and histological examination).
      
Biospecimen description: 
      Colonic washing samples from 4 specific segments (cecum, transverse, proximal sigmoid and
      rectum) of every subject will be retained for 16S rRNA analysis for the study purpose, and
      more samples will be retained for further analysis later on.

      Samples from the unified colonic segments (each collected as 15 cc in each of four 20 cc
      tubes and stored in -70 Freezer) were completely recruited from 50 normal subjects, 50 IBS,
      50 UC, 46 CD and only 29 CRC.

      One to two tubes from each segment sample of all subjects have been utilized for DNA
      extraction and accepted if identified sufficient according to lab. Hence submitted for 16S
      rRNA analysis. Currently we are waiting for intra-cohort comparison of samples, then
      inter-cohort comparisons among different study groups will be done.
    
Keywords: IBS; UC; CD; CRAP/CRC