The regulatory role of anti-sigma factor, RsbW, in Clostridioides difficile stress response, persistence and infection

Abstract

SummaryThe anaerobic pathogen Clostridioides difficile, a primary cause of antibiotic-associated diarrhoea, faces a variety of stresses in the environment and in the mammalian gut. To cope with environmental stresses, it uses the alternative sigma factor B ({sigma}B) to modulate gene transcription, which is regulated by an anti-sigma factor, RsbW. To understand the role of RsbW in C. difficile physiology, a rsbW mutant ({Delta}rsbW) where {sigma}B is always on, was generated. {Delta}rsbW did not have deleterious fitness defects but tolerated acidic environments and detoxified reactive oxygen and nitrogen species better. {Delta}rsbW was defective in spore and biofilm formation, adhered better to human gut epithelia and was less virulent in a Galleria mellonella infection model. A transcriptomic analysis to understand this unique phenotype showed a change in expression of some {sigma}B-controlled genes along with several non-{sigma}B controlled genes. Interestingly, sinRR locus that encodes a pleiotropic regulator, was highly upregulated in {Delta}rsbW indicating a potential indirect role for {sigma}B or RsbW in control of sinRR. Furthermore, the unexpected lower intracellular levels of {sigma}B observed suggest post translational control mechanisms. Our study thus provides new insight into the regulatory role of RsbW and the complexity of regulatory networks in C. difficile. ImportancePathogens, like C. difficile, face a range of stresses in the environment and within the host. Alternate transcriptional factors such as sigma factor B ({sigma}B) enables the bacterium to respond quickly to different types of stresses and are conserved across bacteria. Anti-sigma factors like RsbW control the activation of genes via these pathways. Such transcriptional control systems provide pathogens like C. difficile a route to tolerance and detoxification of harmful compounds. In this study we investigate the role of RsbW in C. difficile physiology. We demonstrate distinctive phenotypes for a rsbW mutant in growth, persistence and virulence. Our data suggest new {sigma}B regulatory circuits in C. difficile. Understanding bacterial responses to external stress is key to designing better strategies to combat this highly resilient bacterial pathogen.