The human intestine is an organ with a unique anatomy, physiology, and microflora. The features of the human gut, together with the presence of various products of food digestion, result in a large variety of ecological niches for the gut microorganisms. The human intestinal microbiota includes up to 100 trillion microbial cells belonging to many hundreds of species. This microbiota is metabolically active and plays a significant role in host physiology, health, and disease. The relationship between the human intestinal microbiota and human health has been intensively studied but previous analysis has mostly focused on the taxonomical diversity of the microbiota.The proposed project aims to develop a fundamentally new approach to use the composition and abundance of specific genes and pathways encoded by the human gut microbiome (HGM) as diagnostic tool. This approach is based on the analysis of the abundance of the specific genes, which can be markers of the host health-state, rather than the taxonomical composition of the HGM alone. As the disease-association of microbial genes is generally not known, this project proposes a workflow to identify such genes. The three considered diseases are Parkinson’s disease, which affects >4 million people world wide and an estimated 1000 individuals in Luxembourg, cardiovascular disease with 17.5 million annual deaths worldwide, and colon cancer with ~100,000 new cases in 2016 in the USA alone. The proposed workflow consists of the following five steps: 1. Eight metabolic pathways relevant for the three human disease will be considered, including respiration, the biosynthesis of indoles, quinones, and short-chain fatty acids, the utilization of ethanolamine and propanediol, and the degradation of betaine, choline, and bile acids. These pathways will be reconstructed in 321 HGM genomes using state-of-the-art comparative genomics methods. 2. We will expand existing genome-scale metabolic reconstructions for the 321 microbes by these disease-relevant pathways. 3. Metagenomic data sets for these three diseases will be retrieved and mapped onto the expanded metabolic reconstructions. 4. We will identify pathways that are either over- or underrepresented each disease metagenome when compared with the healthy control. These pathways are deemed disease-associated pathways. 5. The corresponding genes, encoding the enzymes of the pathways, will be tested for their suitability as disease-marker by excluding those genes that have no homologs in other pathways and that non-orthologous displacements are absent. Genes matching these criteria are considered disease-specific microbial markers in human fecal metagenomes for the corresponding genes.This project will yield a comprehensive and accurate mapping of at least eight metabolic pathways reconstructed for 321 HGM genomes. This project will demonstrate the association of various metabolic pathways with the three considered diseases. The proposed project will provide a milestone towards the HGM-based diagnosis using the over- or underexpression of disease-marker genes in an individual’s metagenome as a predictor for a given disease. As such, the project is likely to result in patentable disease-marker genes. The project results will also improve our understanding of the bacterial biochemistry and transport, biochemical interactions in the HGM and their role in human physiology.This project has been submitted for the CORE Junior program.