Spore formation by Clostridium difficile is a significant obstacle to overcoming hospital-acquired C. difficile-associated disease. Spores are resistant heat, radiation, chemicals, and antibiotics, making contaminated environment difficult clean. To cause disease, however, spores must germinate grow out as vegetative cells. The germination of has not been examined in detail. In an effort understand the spores, we characterized response bile. We found that cholate derivatives amino acid...

ABSTRACT To cause disease, Clostridium difficile spores must germinate in the host gastrointestinal tract. Germination is initiated upon exposure to glycine and certain bile acids, e.g., taurocholate. Chenodeoxycholate, another acid, inhibits taurocholate-mediated germination. By applying Michaelis-Menten kinetic analysis C. spore germination, we found that chenodeoxycholate a competitive inhibitor of germination appears interact with greater apparent affinity than does We also report...

Clostridium difficile spores must germinate in vivo to become actively growing bacteria order produce the toxins that are necessary for disease. C. vitro response certain bile acids and glycine. In other sporulating bacteria, proteins embedded within inner membrane of spore sense presence germinants trigger release Ca++-dipicolinic acid (Ca++-DPA) from core subsequent hydrolysis cortex, a specialized peptidoglycan. Based upon homology searches known germinant receptors spore-forming likely...

ABSTRACT Treatment with antibiotics is a major risk factor for Clostridioides difficile infection, likely due to depletion of the gastrointestinal microbiota. Two microbiota-mediated mechanisms thought limit C. colonization include conversion conjugated primary bile salts into secondary toxic growth and competition between microbiota limiting nutrients. Using continuous flow model that simulates nutrient conditions distal colon, we investigated how treatment 6 clinically used influenced...

Some cholate derivatives that are normal components of bile can act with glycine to induce the germination Clostridium difficile spores, but at least one component, chenodeoxycholate, does not germination. Here we show chenodeoxycholate inhibits C. spores in response and taurocholate.

Clostridium difficile, a spore-forming bacterium, causes antibiotic-associated diarrhea. In order to produce toxins and cause disease, C. difficile spores must germinate grow out as vegetative cells in the host. Although few compounds capable of germinating vitro have been identified, vivo signal(s) which respond were not previously known. Examination intestinal cecal extracts from untreated antibiotic-treated mice revealed that can stimulate colony formation greater levels. Treatment these...

Clostridium difficile is a Gram-positive, spore-forming, anaerobe and the leading cause of antibiotic-associated diarrhea, pseudomembranous colitis, toxic megacolon. Essential to lifestyle C. ability form metabolically dormant spore, germinate, grow out upon appropriate signals elicit disease with secretion two toxins. To aid in study this organism, unit describes growth maintenance difficile. Included are methods isolate from environmental samples, laboratory medium, produce purify spores.

Abstract Clostridium difficile is a significant concern as nosocomial pathogen, and genetic tools are important when analyzing the physiology of such organisms so that underlying physiology/pathogenesis can be studied. Here, we used TargeTron to investigate role selenoproteins in C . Stickland metabolism found insertion into selD , encoding selenophosphate synthetase essential for specific incorporation selenium selenoproteins, results growth defect global loss incorporation. However,...

ABSTRACT Bacterial spore germination is a process whereby dormant returns to active, vegetative growth, and this has largely been studied in the model organism Bacillus subtilis . In B. , initiation of germinant receptor-mediated divided into two genetically separable stages. Stage I characterized by release dipicolinic acid (DPA) from core. II cortex degradation, stage activated DPA released during I. Thus, precedes hydrolysis germination. Here, we investigated timing Clostridium difficile...

Clostridioides difficile infections occur upon ecological / metabolic disruptions to the normal colonic microbiota, commonly due broad-spectrum antibiotic use. Metabolism of bile acids through a 7α-dehydroxylation pathway found in select members healthy microbiota is regarded be protective mechanism by which C. excluded. These 7α-dehydroxylated secondary are highly toxic vegetative growth, and treatment abolishes bacteria that perform this metabolism. However, data supports hypothesis...

Historically, mutagenesis in the non-model enteropathogenic bacterium Clostridioides difficile has been challenging. Developing a versatile and reliable method of generating targeted mutations C. is important to further our understanding its pathogenesis. Some most common systems rely on allelic exchange mediated by either uracil auxotrophy combined with toxic precursor, toxin/anti-toxin system, group II introns, or CRISPR/Cas mutagenesis. However, each these methods suffers from own issues....

The current epidemic of infections caused by antibiotic-resistant Gram-positive bacteria requires the discovery new drug targets and development therapeutics. Lipoteichoic acid (LTA), a cell wall polymer bacteria, consists 1,3-polyglycerol-phosphate linked to glycolipid. LTA synthase (LtaS) polymerizes polyglycerol-phosphate from phosphatidylglycerol, reaction that is essential for growth bacteria. We screened small molecule libraries compounds inhibiting Staphylococcus aureus but not...

ABSTRACT Clostridium difficile is a Gram‐positive, spore forming, anaerobic, intestinal bacterium and the most common cause of antibiotic‐associated colitis. For many years this organism was considered genetically intractable, but in past 10 years, multiple methods have been developed or adapted for genetic manipulation C. . This unit describes molecular techniques used modification organism, including gene disruption, complementation, plasmid introduction integration, cross‐species conjugations.

Clostridium difficile is a Gram-positive spore-forming pathogen and leading cause of nosocomial diarrhea. C. infections are transmitted when ingested spores germinate in the gastrointestinal tract transform into vegetative cells. Germination begins germinant receptor CspC detects bile salts gut. subtilisin-like serine pseudoprotease that activates related CspB protease through an unknown mechanism. Activated cleaves pro-SleC zymogen, which allows activated SleC cortex hydrolase to degrade...

The nosocomial pathogen Clostridioides difficile is a burden to the healthcare system. Gut microbiome disruption, most commonly by broad-spectrum antibiotic treatment, well established generate state that susceptible CDI. A variety of metabolites produced host and/or gut microbiota have been shown interact with C. difficile. Certain bile acids promote/inhibit germination while other cholesterol-derived compounds and amino used in Stickland metabolic pathway affect growth CDI colonization....

The small acid-soluble proteins are found in all endospore-forming organisms and a major component of spores. Through their DNA binding capabilities, the SASPs shield from outside insults (e.g., UV genotoxic chemicals). absence results spores with reduced viability when exposed to light and, at least one case, inability complete sporulation. While have been characterized for decades, some evidence suggests that using newer technologies revisit roles could reveal novel functions spore regulation.

Clostridioides difficile is a pathogen whose transmission relies on the formation of dormant endospores. Spores are highly resilient forms bacteria that resist environmental and chemical insults. In recent work, we found C. SspA SspB, two small acid-soluble proteins (SASPs), protect spores from UV damage and, interestingly, necessary for mature spores. Here, build upon this finding show sspA sspB required spore cortex layer. Moreover, using an EMS mutagenesis selection strategy, identified...

Infections caused by Clostridium difficile have increased steadily over the past several years. While studies on C. virulence and physiology been hindered, in past, lack of genetic approaches suitable animal models, newly developed technologies models allow these processes to be studied detail. One such advance is generation a mouse-model infection. The development this system major step forward analyzing requirements for colonization important, it equally as important understanding what...

ABSTRACT Fidaxomicin (FDX) is approved to treat Clostridium difficile -associated diarrhea and superior vancomycin in providing a sustained clinical response (cure without recurrence the subsequent 25 days). The mechanism(s) behind low rate of FDX-treated patients could be multifactorial. Here, we tested effects FDX, its metabolite OP-1118, on spore germination determined that none affected initiation but all inhibited outgrowth vegetative cells from germinated spores.

ABSTRACT Clostridium difficile spore germination is essential for colonization and disease. The signals that initiate C. are a combination of taurocholic acid (a bile acid) glycine. Interestingly, the chenodeoxycholic class (CDCA) acids competitively inhibit acid-mediated germination, suggesting compounds could be developed into drugs prophylactically prevent infection or reduce recurring However, recent report called question utility such strategy to by describing strains germinated in...

Classically, dormant endospores are defined by their resistance properties, particularly to heat. Much of the heat is due large amount dipicolinic acid (DPA) stored within spore core. During germination, DPA released and allows for rehydration otherwise-dehydrated In Bacillus subtilis, 7 proteins encoded spoVA operon important release. These receive a signal from activated germinant receptor release DPA. This activates cortex lytic enzyme CwlJ, degradation begins. Clostridium difficile,...