The development of Clostridium difficile genetic systems

Minton, Nigel P.; Carter, Glen P.; Herbert, Mike; O'Keeffe, Triona; Purdy, Des; Elmore, Michael; Ostrowski, Anna; Pennington, Oliver; Davis, Ian J.

doi:10.1016/j.anaerobe.2003.11.003

Cited by 17 publications

(12 citation statements)

References 48 publications

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“…One possible alternative was to use an antisense RNA strategy. Such an approach was first exemplified with C. botulinum (16) and has since been used extensively to modulate gene function in C. acetobutylicum (8,9,(34)(35)(36) and, to a lesser extent, in C. perfringens (26) and C. difficile (17). Thus, once the sequences for the two C. sporogenes agr loci had been obtained, this method was used to investigate their functions.…”

Section: Resultsmentioning

confidence: 99%

Regulation of Neurotoxin Production and Sporulation by a Putative agrBD Signaling System in Proteolytic Clostridium botulinum

Cooksley

Davis

Winzer

et al. 2010

Appl Environ Microbiol

115

104

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A significant number of genome sequences of Clostridium botulinum and related species have now been determined. In silico analysis of these data revealed the presence of two distinct agr loci (agr-1 and agr-2) in all group I strains, each encoding putative proteins with similarity to AgrB and AgrD of the well-studied Staphylococcus aureus agr quorum sensing system. In S. aureus, a small diffusible autoinducing peptide is generated from AgrD in a membrane-located processing event that requires AgrB. Here the characterization of both agr loci in the group I strain C. botulinum ATCC 3502 and of their homologues in a close relative, Clostridium sporogenes NCIMB 10696, is reported. In C. sporogenes NCIMB 10696, agr-1 and agr-2 appear to form transcriptional units that consist of agrB, agrD, and flanking genes of unknown function. Several of these flanking genes are conserved in Clostridium perfringens. In agreement with their proposed role in quorum sensing, both loci were maximally expressed during late-exponential-phase growth. Modulation of agrB expression in C. sporogenes was achieved using antisense RNA, whereas in C. botulinum, insertional agrD mutants were generated using ClosTron technology. In comparison to the wild-type strains, these strains exhibited drastically reduced sporulation and, for C. botulinum, also reduced production of neurotoxin, suggesting that both phenotypes are controlled by quorum sensing. Interestingly, while agr-1 appeared to control sporulation, agr-2 appeared to regulate neurotoxin formation.

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Section: Resultsmentioning

confidence: 99%

Regulation of Neurotoxin Production and Sporulation by a Putative agrBD Signaling System in Proteolytic Clostridium botulinum

Cooksley

Davis

Winzer

et al. 2010

Appl Environ Microbiol

115

104

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“…In the early 2000s, an increase in severe cases of CDI was noted in Canada, the United States and Europe 6 , which was attributed to the emergence of certain epidemic types of C. difficile 7,8 . The first complete genome sequence for C. difficile 9 , together with the development of tools for the genetic manipulation of C. difficile 10,11 , has greatly stimulated research on the bacterium. C. difficile is now recognized as the leading cause of health-care-associated infective diarrhoea and is increasingly being linked to community-acquired cases of colitis 12 .…”

mentioning

confidence: 99%

Clostridium difficile infection

Smits

Lyras

Lacy

et al. 2016

Nat Rev Dis Primers

699

674

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Infection of the colon with the Gram-positive bacterium Clostridium difficile is potentially life threatening, especially in elderly people and in patients who have dysbiosis of the gut microbiota following antimicrobial drug exposure. C. difficile is the leading cause of health-care-associated infective diarrhoea. The life cycle of C. difficile is influenced by antimicrobial agents, the host immune system, and the host microbiota and its associated metabolites. The primary mediators of inflammation in C. difficile infection (CDI) are large clostridial toxins, toxin A (TcdA) and toxin B (TcdB), and, in some bacterial strains, the binary toxin CDT. The toxins trigger a complex cascade of host cellular responses to cause diarrhoea, inflammation and tissue necrosis — the major symptoms of CDI. The factors responsible for the epidemic of some C. difficile strains are poorly understood. Recurrent infections are common and can be debilitating. Toxin detection for diagnosis is important for accurate epidemiological study, and for optimal management and prevention strategies. Infections are commonly treated with specific antimicrobial agents, but faecal microbiota transplants have shown promise for recurrent infections. Future biotherapies for C. difficile infections are likely to involve defined combinations of key gut microbiota.

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“…Due to the scarcity of genetic tools, many of the metabolic capabilities encoded by C. difficile remain poorly understood (19). This has precluded the use of molecular microbiology to identify putative germination receptors encoded by C. difficile.…”

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confidence: 99%

Mapping Interactions between Germinants and Clostridium difficile Spores

2011

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Germination of Clostridium difficile spores is the first required step in establishing C. difficile-associated disease (CDAD). Taurocholate (a bile salt) and glycine (an amino acid) have been shown to be important germinants of C. difficile spores. In the present study, we tested a series of glycine and taurocholate analogs for the ability to induce or inhibit C. difficile spore germination. Testing of glycine analogs revealed that both the carboxy and amino groups are important epitopes for recognition and that the glycine binding site can accommodate compounds with more widely separated termini. The C. difficile germination machinery also recognizes other hydrophobic amino acids. In general, linear alkyl side chains are better activators of spore germination than their branched analogs. However, L-phenylalanine and L-arginine are also good germinants and are probably recognized by distinct binding sites. Testing of taurocholate analogs revealed that the 12-hydroxyl group of taurocholate is necessary, but not sufficient, to activate spore germination. In contrast, the 6-and 7-hydroxyl groups are required for inhibition of C. difficile spore germination. Similarly, C. difficile spores are able to detect taurocholate analogs with shorter, but not longer, alkyl amino sulfonic acid side chains. Furthermore, the sulfonic acid group can be partially substituted with other acidic groups. Finally, a taurocholate analog with an m-aminobenzenesulfonic acid side chain is a strong inhibitor of C. difficile spore germination. In conclusion, C. difficile spores recognize both amino acids and taurocholate through multiple interactions that are required to bind the germinants and/or activate the germination machinery.

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The development of Clostridium difficile genetic systems

Cited by 17 publications

References 48 publications

Regulation of Neurotoxin Production and Sporulation by a Putative agrBD Signaling System in Proteolytic Clostridium botulinum

Regulation of Neurotoxin Production and Sporulation by a Putative agrBD Signaling System in Proteolytic Clostridium botulinum

Clostridium difficile infection

Mapping Interactions between Germinants and Clostridium difficile Spores

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