An intact S-layer is advantageous to Clostridioides difficile within the host

Area of Science:

• Microbiology
• Pathogenesis
• Molecular Biology

Background:

• Clostridioides difficile causes significant illness and death, particularly in elderly patients receiving antibiotics.
• While toxins are key virulence factors, the role of other components like the S-layer is less understood.

Purpose of the Study:

• To investigate the in vivo essentiality of the C. difficile S-layer for pathogenesis.
• To characterize S-layer variants that emerge during infection.

Main Methods:

• Infection models using an S-layer-null C. difficile strain (FM2.5).
• Recovery and sequencing of S-layer variants (slpA).
• Structural determination of SlpA variants.
• Comparative RNA sequencing (RNA-Seq) of isolates.

Main Results:

• S-layer variants rapidly emerged in vivo, restoring slpA expression and function.
• Two variants, FM2.5varA and FM2.5varB, showed differential virulence.
• Structural changes in SlpA affected lattice assembly and interactions.
• RNA-Seq revealed distinct gene expression profiles correlating with variant virulence and disease severity.

Conclusions:

• The S-layer is essential for C. difficile pathogenesis and disease severity in vivo.
• S-layer structure and assembly influence virulence.
• Gene expression patterns, including toxin and cell wall genes, are modulated by S-layer status and correlate with disease outcomes.