C. difficile is a gram positive spore forming anaerobic bacterium. C. difficile infection (CDI) mainly develops when a hospitalized patient with disrupted gut microflora due to antibiotic treatment is exposed to C. difficile spores. Other risk factors for CDI include anti-ulcer medications, longer hospital stay, patient age and severe underlying illness. In this study, we proposed to develop colon stem cell organoids that are more closely mimic natural tissue and organ responses that primary cell cultures or available animal models. The cells and the extracellular matrix that they synthesize and secrete in three dimensions (3D) are flexible and pliable like natural tissues. These cell-to-cell interactions in 3D cell culture also include gap junctions that enable cells to communicate with each other via exchange of ions, small molecules and even electrical currents. Therefore, colon stem cell organoids mimic in vivo animal experiments and can be used to study the interaction of C. difficile virulence factors and the host.