Metabolic plasticity enables Clostridioides difficile growth in coculture with neutrophils and other mammalian cells.

OBJECTIVES

Clostridioides difficile infection (CDI) is a significant healthcare burden, characterized by severe colitis and high recurrence rates. Although neutrophils migrate rapidly to the colon infected with C. difficile, it does not lead to pathogen clearance. Understanding how pathogen survives within the host remains a knowledge gap. This study investigates host-pathogen interactions in controlled in vitro settings.

METHODS

Multiple strains of C. difficile (M7404, VPI10463, R20291, and clinical isolates) were co-cultured with neutrophils, macrophages, and intestinal epithelial cells (IECs) under anaerobic conditions, and bacterial growth was assessed by optical density and micro-replicator-mediated colony assays. Microbial transcriptomics and pathway enrichment analyses were performed 18 h after M7404-neutrophil coculture to understand the metabolic adaptations.

RESULTS

Neutrophils significantly accelerated the growth of C. difficile in anaerobic co-culture. Transcriptomic analysis revealed extensive metabolic reprogramming in C. difficile, including upregulation of oxidative phosphorylation, carbon fixation, and fermentation pathways, suggesting that the bacterium exploits host-derived nutrients to its advantage. Further highlighting the microbial metabolic switch towards utilizing host-derived nutrients, we observed the upregulation of glycerol kinase and the ethanolamine utilization protein. The downregulation of TCA cycle enzymes and upregulation of catalases suggested a shift away from oxidative metabolism and an effort to mitigate neutrophil-induced stress. C. difficile bacterium underwent similar transcriptional programming in coculture with macrophages and IECs.

CONCLUSIONS

The transcriptional regulation of C. difficile metabolic genes in the presence of mammalian cells reveals a key virulence trait that enables pathogen persistence within a host. This study also highlights a paradoxical role of neutrophils in CDI, where it's presence may inadvertently enhance pathogen survival. Targeting these metabolic interactions could lead to novel therapeutic strategies for mitigating CDI severity.