The Immune Protein Calprotectin Impacts Clostridioides difficile Metabolism through Zinc Limitation.

The intestines house a diverse microbiota that must compete for nutrients to survive, but the specific limiting nutrients that control pathogen colonization are not clearly defined. colonization typically requires prior disruption of the microbiota, suggesting that outcompeting commensals for resources is critical to establishing infection (CDI). The immune protein calprotectin (CP) is released into the gut lumen during CDI to chelate zinc (Zn) and other essential nutrient metals. Yet, the impact of Zn limitation on colonization is unknown. To define responses to Zn limitation, we performed RNA sequencing on exposed to CP. In medium containing CP, upregulated genes involved in metal homeostasis and amino acid metabolism. To identify CP-responsive genes important during infection, we measured the abundance of select transcripts in a mouse CDI model relative to expression Gene transcripts involved in selenium (Se)-dependent proline fermentation increased during infection and in response to CP. Increased proline fermentation gene transcription was dependent on CP Zn binding and proline availability, yet proline fermentation was only enhanced when Se was supplemented. CP-deficient mice could not restrain proline fermentation-dependent growth, suggesting that CP-mediated Zn sequestration along with limited Se restricts proline fermentation. Overall, these results highlight how colonization depends on the availability of multiple nutrients whose abundances are dynamically influenced by the host response. infection (CDI) is the leading cause of postantibiotic nosocomial infection. Antibiotic therapy can be successful, yet up to one-third of individuals suffer from recurrent infections. Understanding the mechanisms controlling colonization is paramount in designing novel treatments for primary and recurrent CDI. Here, we found that limiting nutrients control metabolism during CDI and influence overall pathogen fitness. Specifically, the immune protein CP limits Zn availability and increases transcription of genes necessary for proline fermentation. Paradoxically, this leads to reduced proline fermentation. This reduced fermentation is due to limited availability of another nutrient required for proline fermentation, Se. Therefore, CP-mediated Zn limitation combined with low Se levels overall reduce fitness in the intestines. These results emphasize the complexities of how nutrient availability influences colonization and provide insight into critical metabolic processes that drive the pathogen's growth.