The Clostridioides difficile taxonomy is characterised by major taxonomic incoherence and clear species boundaries.

Immunosuppressed patients demonstrate impaired immune responses to Clostridioides difficile treatment compared with patients who are not immunosuppressed.

The formation of dormant spores is essential for the anaerobic pathogen Clostridioides difficile to survive outside of the host gastrointestinal tract. The regulatory pathways and environmental signals that initiate C. difficile spore formation within the host are not well understood. One bacterial second messenger signaling molecule, cyclic diguanylate (c-di-GMP), modulates several physiological processes important for C. difficile pathogenesis and colonization, but the impact of c-di-GMP on sporulation is unknown. In this study, we investigated the contribution of c-di-GMP to C. difficile sporulation. Overexpression of a gene encoding a diguanylate cyclase, dccA , decreased sporulation frequency and early sporulation gene transcription in both the epidemic R20291 and historical 630Δ erm strains. Expression of a dccA allele encoding a catalytically inactive DccA that is unable to synthesize c-di-GMP no longer inhibited sporulation, indicating that the accumulation of intracellular c-di-GMP reduces C. difficile sporulation. A null mutation in dccA slightly increased sporulation in R20291 and slightly decreased sporulation in 630Δ erm , suggesting that DccA may contribute to the intracellular pool of c-di-GMP in a strain-dependent manner. However, these data were highly variable, underscoring the complex regulation involved in modulating intracellular c-di-GMP concentrations. Finally, overexpression of dccA in known sporulation mutants revealed that c-di-GMP is likely signaling through an unidentified regulatory pathway to control early sporulation events in C. difficile . C-di-GMP-dependent regulation of C. difficile sporulation may represent an unexplored avenue of potential environmental and intracellular signaling that contributes to the complex regulation of sporulation initiation. IMPORTANCE Many bacterial organisms utilize the small signaling molecule cyclic diguanylate (c-di-GMP) to regulate important physiological processes, including motility, toxin production, biofilm formation, and colonization. C-di-GMP inhibits motility and toxin production and promotes biofilm formation and colonization in the anaerobic, gastrointestinal pathogen Clostridioides difficile . However, the impact of c-di-GMP on C. difficile spore formation, a critical step in this pathogen’s life cycle, is unknown. Here, we demonstrate that c-di-GMP negatively impacts sporulation in two clinically relevant C. difficile strains, the epidemic R20291 and the historical 630Δ erm . The pathway through which c-di-GMP controls sporulation was investigated, and our results suggest that c-di-GMP is likely signaling through an unidentified regulatory pathway to control C. difficile sporulation. This work implicates c-di-GMP metabolism as a potential mechanism to integrate environmental and intracellular cues through c-di-GMP levels to influence C. difficile sporulation.

Abstract. Along with the rise in modern chronic diseases, ranging from diabetes to asthma, there are challenges posed by increasing antibiotic resistance, which

The anaerobic pathogen Clostridioides difficile is perfectly equipped to survive and persist inside the mammalian intestine. When facing unfavorable conditions C. difficile is able to form highly resistant endospores. Likewise, biofilms are currently discussed as form of persistence. H …

Cellobiose metabolism is linked to the virulence properties in numerous bacterial pathogens. Here, we characterized a putative cellobiose PTS operon of Clostridiodes difficile to investigate the role of cellobiose metabolism in C. difficile pathogenesis. Our gene knockout experiments demonstrated that the putative cellobiose operon enables uptake of cellobiose into C. difficile and allows growth when cellobiose is provided as the sole carbon source in minimal medium. Additionally, using reporter gene fusion assays and DNA pull-down experiments, we show that its transcription is regulated by CelR, a novel transcriptional repressor protein, which directly binds to the upstream region of the cellobiose operon to control its expression. We have also identified cellobiose metabolism to play a significant role in C. difficile physiology as observed by the reduction of sporulation efficiency when cellobiose uptake was compromised in the mutant strain. In corroboration to in vitro study findings, our in vivo hamster challenge experiment showed a significant reduction of pathogenicity by the cellobiose mutant strain in both the primary and the recurrent infection model- substantiating the role of cellobiose metabolism in C. difficile pathogenesis. ### Competing Interest Statement The authors have declared no competing interest.

Clostridioides difficile BI/NAP1/ribotype 027 is an epidemic hypervirulent strain found worldwide, including in Latin America. We examined the genomes and exoproteomes of two multilocus sequence type (MLST) clade 2 C. difficile strains considered hypervirulent: ICC-45 (ribotype SLO231/UK[CE]821), is …

This study found that ongoing exposure to PPIs was associated with a doubled risk of community-acquired Clostridium difficile infection. The risk was attenuated

C.difficile infection (CDI) is not a merely "gut-confined" disease as toxemia could drive the development of CDI-related extra-intestinal effects. These effects could explain the high CDI-associated mortality, not just justified by diarrhea and dehydration. Here, the extra-intestinal e …

Immune checkpoint inhibitors (ICIs) are monoclonal antibodies that block immune inhibitory pathways. Administration of ICIs augments T cell-mediated immune responses against tumor, resulting in improved overall survival in cancer patients. It has emerged that the intestinal microbiome can modulate r …

Researchers at UCSF have found that extreme caloric restriction diets alter the microbiome in ways that could help with weight loss but might also result in an increased population of Clostridiodes difficile, a pathogenic bacterium that can lead to severe diarrhea and colitis.

The overall 90-day cumulative incidences of a first CDI episode was 1.9%.

On the HCPLive C. difficile condition center page, resources on the topics of medical news and expert insight into Clostridium difficile can be found. Content includes articles, interviews, videos, podcasts, and breaking news on C. diff research, treatment, and drug development.

Severe caloric restriction in humans leads to reversible changes in the gut microbiota that promote weight loss and the expansion of an enteric pathogen in mice.

The phosphorous balance is clinically important in increasing the long-term outcomes and preventing complications of end-stage renal disease. Sevelamer is a phosphate binder used widely to regulate hyperphosphatemia. On the other hand, gastrointestinal side effects increase with increasing sevelamer …

Researchers were able to show for the first time that a very low calorie diet significantly alters the composition of the microbiota present in the human gut. The researchers report that dieting results in an increase of specific bacteria - notably Clostridioides difficile, which is associated with antibiotic-induced diarrhea and colitis.

Antimicrobial resistance has become a worldwide problem due to its excessive increase in recent years. The aim of the present review was to bring together data from different articles describing the levels of antimicrobial resistance in the most common gastrointestinal infections reported across the …

Spore-forming pathogens like Clostridioides difficile depend on germination to initiate infection. Spore germination depends on the degradation of the protective spore peptidoglycan layer known as the spore cortex. Cortex degradation is mediated by enzymes that recognize the spore-specific peptidoglycan modification, muramic-∂-lactam (MAL). In C. difficile, MAL synthesis depends on the activity of the CwlD amidase and the GerS lipoprotein, which directly binds CwlD. To gain insight into how GerS regulates CwlD activity, we solved the crystal structure of the CwlD:GerS complex. In this structure, a GerS homodimer is bound to two CwlD monomers such that the CwlD active sites are exposed. Although CwlD structurally resembles amidase\_3 family members, we found that CwlD does not bind zinc stably on its own, unlike previously characterized amidase\_3 enzymes. Instead, GerS binding to CwlD promotes CwlD binding to zinc, which is required for its catalytic mechanism. Thus, in determining the first structure of an amidase bound to its regulator, we reveal stabilization of zinc co-factor binding as a novel mechanism for regulating bacterial amidase activity. Our results further suggest that allosteric regulation by binding partners may be a more widespread mode for regulating bacterial amidase activity than previously thought. ### Competing Interest Statement AS is a consultant and holds shares in a diagnostic start-up company, BioVector, Inc.

Clostridioides difficile infections are associated with gut microbiome dysbiosis and are the leading cause of hospital-acquired diarrhoea. The infectious process is strongly influenced by the microbiota and successful infection relies on the absence of specific microbiota-produced metabolites. Deoxy …

AbstractBackground. Clostridioides difficile was listed as an urgent antimicrobial resistance (AMR) threat in a report by the CDC in 2019. AMR drives the evolut

Clostridioides difficile (C. difficile) is classified as an urgent antibiotic resistance threat by the CDC. The 3-D structure shows how a key C. difficile toxin, TcdB (grey surface model), engages the human receptor CSPG4 (shown in green) for cell entry, and how an FDA-approved therapeutic antibody bezlotoxumab (shown in blue and purple) recognize two epitopes on TcdB. Some C. difficile hypervirulent strains evolve mutations in the bezlotoxumab-binding sites (blue and purple surface, while the mutated amino acids are colored red) that weaken the antibody potency. In contrast, the CSPG4-binding site on TcdB (gold surface) is highly conserved, suggesting a strategy to develop broad-spectrum therapeutics against TcdB.

Clostridioides difficile (C. difficile) infection is the most common cause of healthcare-associated infection and an important cause of morbidity and mortality among hospitalized patients. A comprehensive understanding of C. difficile infection (CDI) pathogenesis i …

Researchers observed the structure of a lethal toxin produced by certain strains of Clostridium difficile bacteria.

Clostridioides difficile is a noteworthy pathogen in patients with inflammatory bowel disease (IBD). Patients with IBD who develop concurrent C. difficile infection (CDI) experience increased morbidity and mortality. IBD is associated with intestinal inflammation and alterations of the gut microbiot …

There was no significant difference in performance between the model with only bile acids compared to models with bile acids and clinical variables.