Probing Clostridium difficile infection in innovative human gut cellular models

Interactions of anaerobic gut bacteria, such as Clostridium difficile , with the intestinal mucosa have been poorly studied due to challenges in culturing anaerobes with the oxygen-requiring gut epithelium. Although gut colonization by C. difficile is a key determinant of disease outcome, precise mechanisms of mucosal attachment and spread remain unclear. Here, using human gut epithelial monolayers co-cultured within dual environment chambers, we demonstrate that C. difficile adhesion to gut epithelial cells is accompanied by a gradual increase in bacterial numbers. Prolonged infection causes redistribution of actin and loss of epithelial integrity, accompanied by production of C. difficile spores, toxins and bacterial filaments. This 2-D dual chamber system was used to examine C. difficile interactions with the commensal Bacteroides dorei , and interestingly, C. difficile growth is significantly reduced in presence of B. dorei . Furthermore, in novel multilayer and 3-D gut models containing a myofibroblast layer, C. difficile adheres more efficiently to epithelial cells, as compared to the 2-D model, leading to a quicker destruction of the epithelium. Our study describes new controlled environment human gut models that enable host-anaerobe and pathogen-commensal interaction studies in vitro .