The Gut Microbiome and Butyrate Differentiate Clostridioides difficile Colonization and Infection in Children

Nicholson, M.; Ma, S.; Stricland, B. A.; Cecala, M.; Zhang, L.; Reasoner, S.; Guiberson, E.; Munneke, M.; Shilts, M. H.; Skaar, E. P.; Das, S. R.

doi:10.1101/2025.04.16.648821

bioRxiv · 10.1101/2025.04.16.648821

The Gut Microbiome and Butyrate Differentiate Clostridioides difficile Colonization and Infection in Children

Nicholson, M.·Ma, S.·Stricland, B. A.·Cecala, M.·Zhang, L.·Reasoner, S.·Guiberson, E.·Munneke, M.·Shilts, M. H.·Skaar, E. P.·Das, S. R.

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Abstract

Background and AimsSymptomatic Clostridioides difficile infection (CDI) can cause significant morbidity and mortality. Conversely, patients can be colonized with toxigenic C. difficile in the absence of symptoms, termed asymptomatic colonization. We previously demonstrated that the presence and function of C. difficile toxins do not differentiate between asymptomatic colonization and CDI in children, suggesting the influence of other factors. This study aimed to interrogate the intestinal microbiome and butyrate in stool samples from children with CDI and asymptomatic colonization. MethodsDesign: Case-control study Setting: Tertiary care childrens hospital Participants and measures: Asymptomatic children had stool tested for C. difficile by nucleic-acid amplification-based testing (NAAT) and were considered colonized if positive (N=50). Residual stool was also obtained from symptomatic children who tested positive for C. difficile by NAAT (N=55). The microbiome was assessed via 16S rRNA sequencing and butyrate via liquid chromatography-mass spectrometry. ResultsCompared to clinical co-variates and comorbidities, C. difficile symptom status (i.e., asymptomatic colonization versus symptomatic CDI) demonstrated the strongest differential abundance association on gut microbes. Symptomatic CDI was associated with increased abundance of Escherichia/Shigella (Benjamini-Hochberg adjusted q=3.94x10-5), Haemophilus (q=0.022), and Gemella (q=0.085), and depleted abundance of gut commensals such as Faecalibacterium (q=0.041), Blautia (q=0.041), and Bifidobacterium (q=0.063). We also observed depletion in the abundance of microbial butyrate producers and fecal butyrate in participants with symptomatic CDI versus asymptomatic colonization. ConclusionThe gut microbiota and butyrate differ between participants with asymptomatic C. difficile colonization and symptomatic CDI, suggesting their potential role in symptom development.

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Nicholson, M., Ma, S., Stricland, B. A., Cecala, M., Zhang, L., Reasoner, S., Guiberson, E., Munneke, M., Shilts, M. H., Skaar, E. P., Das, S. R.. 2025-04-16. The Gut Microbiome and Butyrate Differentiate Clostridioides difficile Colonization and Infection in Children. https://doi.org/10.1101/2025.04.16.648821

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