Investigating Clostridioides difficile Genes Important during Zinc Limitation in the Presence of the Intestinal Microbiota
Clostridioides difficile is an intestinal bacterial pathogen that causes severe diarrhea, pseudomembranous colitis, toxic megacolon, and even death. This bacterium currently putting more financial strain on the healthcare system as incidence of C. difficile infection (CDI) and death rates have been on the rise. The intestinal microbiota plays a role in preventing CDI as the microbiota is thought regulate the immune response to CDI and compete with the C. difficile for nutrients, but specific mechanisms of competition have not been fully defined. Zinc is a vital nutrient for C. difficile growth and therefore its ability to cause disease. The immune system exploits the necessity for zinc by producing the metal-chelating protein calprotectin (CP). CP binds zinc from the intestinal environment to starve C. difficile and other bacterial pathogens. It is also possible that representative members of the microbiota compete with C. difficile for zinc. Considering the host-induced zinc limitation and the importance of the microbiota for colonization resistance, we hypothesized that there are specific genes within the C. difficile genome that are important to maintain zinc homeostasis during limitation conditions when the commensals, Bacteroides, are present. To test this hypothesis, we used the next-generation sequencing technique, Transposon Sequencing (Tn-Seq) by competing a C. difficile transposon mutant library with Bacteroides thetaiotaomicron and Bacteroides fragilis in zinc deplete and replete conditions. We discovered that a mutation in feoB1, a putative ferrous iron transporter, was selected against during zinc limitation, and we aim to investigate the specific role of this gene during zinc limitation, similar to that experienced in the intestines. Taken together, Tn-Seq can be used to determine genes important for colonization and infection. This improved understanding of mechanisms for colonization and infection in C. difficile could lead to a new generation therapeutics that specifically inhibit these mechanisms.
McNeely, Tess, "Investigating Clostridioides difficile Genes Important during Zinc Limitation in the Presence of the Intestinal Microbiota" (2022). Honors Theses. 66.