Language

English

Publication Date

8-26-2026

Journal

mSphere

DOI

10.1128/msphere.00263-26

PMID

42461050

PMCID

PMC13508786

PubMedCentral® Posted Date

7-16-2026

PubMedCentral® Full Text Version

Post-print

Abstract

Hundreds of ribotypes of the gastrointestinal pathogen Clostridioides difficile have emerged over the last three decades, yet the factors driving their emergence are poorly understood. Recently, there has been an increase in infections caused by PCR ribotype 023 (RT023) strains in Europe. We profiled the growth of seven RT023 strains in 190 unique carbon sources and found they were able to grow in xylitol, a sugar alcohol used as a food additive in humans and animals. Other ribotypes of C. difficile tested (n = 19) displayed little to no growth in 0.5% xylitol and were growth-inhibited in higher concentrations of xylitol. Genome sequencing identified that RT023 strains acquired a putative xylitol dehydrogenase (xdh) gene in a mobile genetic element (MGE) that is absent from other C. difficile ribotypes. We created a deletion of xdh in the RT023 strain PRB1128 and observed poor growth in xylitol, indicating that the xdh is necessary for xylitol utilization. Complementation of the xdh mutant with a plasmid-based inducible copy of the xdh gene restored growth in xylitol. We performed competition assays in minibioreactor arrays (MBRAs) and observed that PRB1128 outcompeted the non-xylitol-utilizing strain CD2015 (RT027) in the presence of xylitol. These data support that the xdh gene within RT023 strains provides a fitness benefit for growth in xylitol. Interestingly, the chromosomal locus where the MGE inserted appears to be a hotspot for genetic insertions across clades of C. difficile. Together, this work improves our understanding of the molecular basis for niche adaptation of C. difficile.

IMPORTANCEGenetic factors aiding in the emergence of the opportunistic pathogen Clostridioides difficile are poorly understood. Infections with clade 3 (PCR ribotype 023) strains causing severe disease have increased since 2008. Here, we show RT023 strains have the unique ability to utilize xylitol, a sugar alcohol used as a food additive in humans and animals, due to the presence of a xylitol dehydrogenase (xdh) gene within a mobile genetic element (MGE). This xylitol utilization ability confers a fitness benefit in competition against other C. difficile ribotypes, as well as in a fecal community in vitro. Research investigating the underlying genetic factors driving the physiology of C. difficile will improve our understanding of colonization and hypervirulence.

Keywords

Clostridioides difficile, Xylitol, Ribotyping, Multigene Family, D-Xylulose Reductase, Humans, Interspersed Repetitive Sequences, Clostridium difficile, xylitol, RT023, xdh, mobile genetic elements

Published Open-Access

yes

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