No abstract

Inflammatory bowel diseases (IBD), such as Crohn’s disease, are chronic, immunologically mediated disorders that have severe medical consequences. The current hypothesis is that these diseases are due to an overly aggressive immune response to a subset of commensal enteric bacteria. Studies to date on IBD have suggested that the disorder may be caused by a combination of bacteria and host susceptibility; however the etiologies of these diseases remain an enigma. In this application, we propose to develop and demonstrate the ability to profile Crohn’s disease at an unprecedented molecular level by elucidation of specific biomarkers (bacterial strains, genes, or proteins) that correlate to disease symptoms. To achieve this goal, we will employ a multidisciplinary approach based on metagenomic and metaproteomic molecular tools to elucidate the composition of the commensal microbiota in monozygotic twins that are either healthy or exhibit Crohn’s disease (for concordant, both are diseased; for discordant, one is healthy and one is diseased). The central hypotheses of this proposal are (1) that specific members and/or functional activities of the gastrointestinal (GI) microbiota differ in patients with Crohn’s disease as compared to healthy individuals, and (2) that it will be possible to elucidate microbial signatures which correlate with the occurrence and progression of this disease by integration of data obtained from 16S rRNA based molecular fingerprinting, metagenomics, and metaproteomics approaches. To address these hypotheses, three specific aims are proposed: 1) Obtain data on community gene content (metagenome) in a subset of healthy twins and twins with Crohn’s Disease to assess potential differences in the metabolic capabilities of the gut microbiota associated with CD, 2) Obtain data on community protein content (metaproteome) in a subset of healthy twins and twins with Crohn’s Disease to assess the state of expressed proteins associated with CD, 3) Apply various statistical clustering and classification methods to correlate/associate microbial community composition, gene and protein content with patient metadata, including metabolite profiles and clinical phenotype. The ultimate goal of these efforts is to identify novel biomarkers for non-invasive diagnostics of CD and to eventually identify drug targets (i.e. bacterial strains) for cure or suppression of disease symptoms.

The human species is dependent for its survival upon the activities of billions of microorganisms that inhabit multiple environmental niches within and on the human body. Our current view of microbial‐host interactions is extremely limited by the fact that the overwhelming majority of microbial species (>99%) resists cultivation in the laboratory. Their identification and characterization requires the use of molecular approaches. The impact of this vast number of uncharacterized microbes on human health and disease is potentially significant and remains to be elucidated. The identification and characterization of these microbial communities will undoubtedly establish links between these microorganisms and disease, their roles in development of the immune system, and their overall impact on human evolution. The Human Microbiome Project (HMP), which is a collection of projects around the world, is just being launched with the ultimate goals being to better understand our microbiota and to ultimately manipulate it so as to optimize its beneficial effects on each individual. In this presentation, I will focus on a number of the conceptual and experimental challenges associated with the HMP, with particular emphasis on the human gastrointestinal tract, since this environment is home to the largest number of our microbial partners. (This work was funded in part by NIH grants R01 DE014868 and P01 DK078669).

Emerging evidence that the gut microbiota may contribute in important ways to human health and disease has led us and others to hypothesize that both symbiotic and pathological relationships between gut microbes and their host may by key contributors to obesity and the metabolic complications of obesity. Our “Thrifty Microbiome Hypothesis” posits that gut microbiota play a key role in human energy homeostasis. Specifically, constituents of the gut microbial community may introduce a survival advantage to its host in times of nutrient scarcity, promoting positive energy balance by increasing efficiency of nutrient absorption and improving metabolic efficiency and energy storage. However, in the presence of excess nutrients, fat accretion and obesity may result, and in genetically predisposed individuals, increased fat mass may result in preferential abdominal obesity, ectopic fat deposition (liver, muscle), and metabolic complications of obesity (insulin resistance, hypertension, hyperlipidemia). Furthermore, in the presence of excess nutrients, a pathological transition of the gut microbial community may occur, causing leakage of bacterial products into the intestinal lymphatics and portal circulation, thereby inducing an inflammatory state, further aggravating metabolic syndrome traits and accelerating atherosclerosis. This pathological transition and the extent to which antimicrobial leakage occurs and causes inflammatory and other maladaptive sequelae of obesity may also be influenced by host factors, including genetics. In the proposed study, we will directly test the Thrifty Mirobiome Hypothesis by performing detailed genomic and functional assessment of gut microbial communities in intensively phenotyped and genotyped human subjects before and after intentional manipulation of the gut microbiome. To address this hypothesis, we will enroll three age- and sex-matched groups from the Old Order Amish: (i) 50 obese subjects (BMI > 30 kg/m2) with one or more dysmetabolic manifestations (malignant obesity), (ii) 50 obese subjects (BMI > 30 kg/m2) without any dysmetabolic manifestations (benign obesity), and (iii) 50 nonobese subjects (BMI < 25 kg/m2) without any features of the metabolic syndrome and characterize the architecture of the gut microbiota from the subjects enrolled in this study by high-throughput sequencing of 16S rRNA genes. These studies will provide a deeper understanding of the role of gut microbes in terms of ‘who’s there?’, ‘what are they doing?’, and ‘how are they influencing host energy homeostasis, obesity and its metabolic complications?’

The genomes of 46 archaeal species have been fully sequenced and published. As of this writing (September, 2007), 60 more are in various stages of progress. Analyses of these genomes are providing many useful insights into the evolution and functioning of diverse archaea, helping to understand the diverse physiological properties of archaea and their relationships to bacteria and eukarya.