# The phi027 bacteriophage influences physiology and virulence of the lysogenic strain of Clostridioides difficile

## TL;DR

A bacteriophage called phi027 affects the behavior and virulence of Clostridioides difficile, a bacterium that causes infections in the gut.

## Contribution

The study shows that curing C. difficile of phi027 reduces its virulence and interaction with human colon cells.

## Key findings

- Removing phi027 from C. difficile reduces sporulation efficiency and adhesion to human colon cells.
- Prophage-free C. difficile produces less TcdB, a toxin linked to cytopathic effects.
- The study highlights the role of bacteriophages in shaping C. difficile virulence.

## Abstract

Clostridioides difficile, the causative agent of C. difficile infections (CDI), can be naturally infected by bacterial viruses known as bacteriophages. All characterized bacteriophages of this bacterium are temperate, meaning that upon infection their genetic material integrates and replicates with host’s genome. Such lysogenic strains can exhibit altered physiology and virulence, which in turn can be an important factor for epidemiology of CDI. In this study we characterized the phiCDKH02 bacteriophage infecting clinical isolates of C. difficile belonging to hypervirulent ribotypes 027 and 176. The bacteriophage was found to be identical to phi027. To get some insight into the role of this bacteriophage in physiology of its host and interaction with human colon cells, we made use of CRISPR-Cpf1 technology to cure the lysogenic C. difficile of the prophage. The prophage-free strain exhibited altered sporulation efficiency, lowered adhesion and decreased cytopathic effects towards human colon cells associated with decreased production of TcdB. These results emphasize importance of prophages in shaping virulence of C. difficile.

The online version contains supplementary material available at 10.1038/s41598-025-04106-0.