PROJECT SUMMARY Microscopic colitis is the most common cause of chronic watery diarrhea in older adults. The symptoms of this disease are debilitating. Patients describe isolation and withdrawal from social life and activities. Treatment is limited to expensive therapies that are sometimes ineffective. Disease relapse is common after medications are stopped. Although the incidence is comparable to inflammatory bowel disease, the disease has received much less research attention. A number of medications (PPIs, NSAIDs, statins, SSRIs) have been linked with microscopic colitis. Although guidelines suggest discontinuing these medications as the initial step in treatment, the evidence supporting this recommendation is weak. The first phase of this study enrolled patients with microscopic colitis and diarrhea controls. Contrary to existing beliefs, there was no association with medications previously linked to microscopic colitis. There was a strong inverse association with obesity and exogenous hormones. Amplicon sequencing by 16S rRNA demonstrated that microbiome alpha-diversity was significantly lower in microscopic colitis cases compared to controls. Several taxa were enriched in cases. The present study is designed to extend and expand on the initial findings. The aims of the proposed case-control study are: 1) To investigate medications, oral contraceptives, postmenopausal hormones and obesity by comparing microscopic colitis cases to two comparison groups – patients referred for colonoscopy for diarrhea and colonoscopy for colorectal cancer screening. 2) To use shotgun metagenomic sequencing to gain insight into biodiversity and function. 3) To use RNA sequencing of mucosal samples from microscopic colitis patients, diarrhea controls and screening controls to reveal genetic signatures associated with microscopic colitis and discover potential druggable disease targets. The study will enroll 150 microscopic colitis cases, 300 diarrhea controls and 300 screening colonoscopy controls. Biopsies from the colon will be used to evaluate adherent bacterial organisms and to conduct RNAseq experiments in 100 microscopic colitis cases, 100 diarrhea controls and 100 screening colonoscopy controls. Structured telephone interviews will obtain detailed dietary, medication and lifestyle information on study subjects. The prospective data collection corrects important limitations of prior research by others. The addition of a second control group could provide more definitive evidence to avoid unnecessarily stopping therapeutically important medications (PPIs, NSAIDs, statins, SSRIs). Shotgun metagenomics could provide information on microbial etiology or functional genes potentially leading to strategies to prevent or treat the disease. The study will improve our understanding of risk factors, set the stage for more scientifically grounded future research, and potentially suggest new interventions for a disease that is currently poorly understood.