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bioRxiv · 10.1101/2024.10.30.621113

Impacts of perR on oxygen sensitivity, gene expression, and murine infection in Clostridioides difficile 630Δerm

Gregory, A. L.·Bussan, H. E.·Topf, M. A.·Hryckowian, A. J.
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Abstract

Clostridioides difficile infection (CDI), characterized by colitis and diarrhea, afflicts approximately half a million people in the United States every year, burdening both individuals and the healthcare system. C. difficile 630{Delta}erm is an erythromycin-sensitive variant of the clinical isolate C. difficile 630 and is commonly used in the C. difficile research community due to its genetic tractability. 630{Delta}erm possesses a point mutation in perR, an autoregulated transcriptional repressor that regulates oxidative stress resistance genes. This point mutation results in a constitutively de-repressed PerR operon in 630{Delta}erm. To address the impacts of perR on phenotypes relevant for oxygen tolerance and relevant to a murine model of CDI, we corrected the point mutant to restore PerR function in 630{Delta}erm (herein, 630{Delta}erm perRWT). We demonstrate that there is no difference in growth between 630{Delta}erm and a 630{Delta}erm perRWT under anaerobic conditions or when exposed to concentrations of O2 that mimic those found near the surface of the colonic epithelium. However, 630{Delta}erm perRWT is more sensitive to ambient oxygen than 630{Delta}erm, which coincides with alterations in expression of a variety of perR-dependent and perR-independent genes. Finally, we show that 630{Delta}erm and 630{Delta}erm perRWT do not differ in their ability to infect and cause disease in a well-established murine model of CDI. Together, these data support the hypothesis that the perR mutation in 630{Delta}erm arose as a result of exposure to ambient oxygen and that the perR mutation in 630{Delta}erm is unlikely to impact CDI-relevant phenotypes in laboratory studies. ImportanceClostridioides difficile is a diarrheal pathogen and a major public health concern. To improve humans understanding of C. difficile, a variety of C. difficile isolates are used in research, including C. difficile 630{Delta}erm. 630{Delta}erm is a derivative of the clinical isolate 630 and is commonly studied because it is genetically manipulable. Previous work showed that a mutation in perR in 630{Delta}erm results in a dysregulated oxidative stress response, but no work has been done to characterize perR-dependent effects on the transcriptome or to determine impacts of perR during infection. Here, we identify transcriptomic differences between 630{Delta}erm and 630{Delta}erm perRWT exposed to ambient oxygen and demonstrate that there is no strain-based difference in burdens in murine C. difficile infection.

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Gregory, A. L., Bussan, H. E., Topf, M. A., Hryckowian, A. J.. 2024-10-31. Impacts of perR on oxygen sensitivity, gene expression, and murine infection in Clostridioides difficile 630Δerm. https://doi.org/10.1101/2024.10.30.621113

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