Development and application of a multiple typing system for Clostridium difficile

Mahony, D. E.; Clow, J; Atkinson, Logan; Vakharia, Narendra; Schlech, WalterF.

doi:10.1128/aem.57.7.1873-1879.1991

Cited by 32 publications

(9 citation statements)

References 31 publications

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“…The various phenotypic typing systems that have been applied to C. difficile isolates have recently been reviewed (35). Antibiotic susceptibility, bacteriocin production and susceptibility patterns, and cytotoxicity all have limited discriminatory power (21,33,38). The use of polyacrylamide gel electrophore-sis (PAGE) to analyze bacterial proteins is somewhat more effective, particularly when coupled with [35S]methionine labelling or immunoblotting (27,35).…”

Section: Discussionmentioning

confidence: 99%

“…dificile and to resolve epidemiologically related cases. The earlier typing systems based on phenotypic traits or on plasmid and phage profiles were insufficient for comprehensive analyses, since phenotypes lack discriminatory power and not all strains are typeable by phage and plasmid studies (10,14,16,21,33,37,38). Subsequently, C. difficile typing based on restriction endonuclease analysis (REA) was shown to be reproducible, highly discriminatory, and universally applicable (8,15,17,20).…”

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confidence: 99%

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Comparison of restriction endonuclease analysis, ribotyping, and pulsed-field gel electrophoresis for molecular differentiation of Clostridium difficile strains

et al. 1994

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A combined clinical and molecular epidemiologic analysis of 46 strains of Clostridium dijfile, including 16 nosocomial isolates from one ward (outbreak ward) plus 17 other nosocomial isolates and 13 communityacquired isolates, was performed. HindIll digests of total cellular DNA were analyzed by restriction enzyme analysis (REA) and ribotyping; SmaI digests were analyzed by pulsed-field gel electrophoresis (PFGE). Isolates were assigned to typing groups on the basis of the profiles detected; isolates with closely related profiles were assigned to subgroups. The 16 isolates from the outbreak ward were resolved by both REA and PFGE into five distinct groups; 13 isolates represented two REA groups and three PFGE groups and two isolates were resolved as distinct groups by both techniques. DNA obtained from one isolate was persistently partially degraded, precluding analysis by PFGE. Seventeen sporadic nosocomial isolates were resolved by REA and PFGE into comparable numbers of groups (i.e., nine groups) and subgroups (i.e., 15 and 14 subgroups, respectively), with two isolates not evaluable by PFGE. The 13 epidemiologically unrelated community-acquired isolates were assigned to 11 groups by REA and to 12 groups by PFGE. Overall, ribotyping identified only nine groups among the 46 isolates. We conclude that REA and PFGE have comparable discriminatory powers for epidemiologic typing of C. difile isolates and that ribotyping is appreciably less discriminatory. For a few isolates, partial DNA degradation prevented analysis by PFGE but not by REA or ribotyping; the cause of the degradation is unknown. Clostridium difficile causes a spectrum of colonic diseases ranging from antibiotic-associated diarrhea to pseudomembranous colitis and produces significant morbidity among hospitalized patients (4). Sporadic cases of C. difficile disease may represent overgrowth of endogenous organisms (7), but nosocomial clusters of cases have been ascribed to interpatient transfer of organisms through contaminated environmental sources as well as by hospital personnel (9, 26). Multiple techniques have been used to identify distinct strains of C.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Comparison of restriction endonuclease analysis, ribotyping, and pulsed-field gel electrophoresis for molecular differentiation of Clostridium difficile strains

et al. 1994

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“…Unfortunately, we were unable to find a suitable host to propagate phiCD211. This was not surprising considering the rather narrow host spectrum observed so far with C. difficile phages (9,18,47,48). This may be due to the presence of wide-spectrum antiphage systems (49), including a functional CRISPR-Cas system (19,50).…”

Section: Discussionmentioning

confidence: 84%

High Prevalence and Genetic Diversity of Large phiCD211 (phiCDIF1296T)-Like Prophages in Clostridioides difficile

Garneau

Sekulović

Dupuy

et al. 2018

Appl Environ Microbiol

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(formerly ) is a pathogenic bacterium displaying great genetic diversity. A significant proportion of this diversity is due to the presence of integrated prophages. Here, we provide an in-depth analysis of phiCD211, also known as phiCDIF1296T, the largest phage identified in so far, with a genome of 131 kbp. It shares morphological and genomic similarity with other large siphophages, like phage 949, infecting , and phage c-st, infecting A PhageTerm analysis indicated the presence of 378-bp direct terminal repeats at the phiCD211 genome termini. Among striking features of phiCD211, the presence of several transposase and integrase genes suggests past recombination events with other mobile genetic elements. Several gene products potentially influence the bacterial lifestyle and fitness, including a putative AcrB/AcrD/AcrF multidrug resistance protein, an EzrA septation ring formation regulator, and a spore protease. We also identified a CRISPR locus and a gene. We screened 2,584 genomes available and detected 149 prophages sharing ≥80% nucleotide identity with phiCD211 (5% prevalence). Overall, phiCD211-like phages were detected in strains corresponding to 21 different multilocus sequence type groups, showing their high prevalence. Comparative genomic analyses revealed the existence of several clusters of highly similar phiCD211-like phages. Of note, large chromosome inversions were observed in some members, as well as multiple gene insertions and module exchanges. This highlights the great plasticity and gene coding potential of the phiCD211/phiCDIF1296T genome. Our analyses also suggest active evolution involving recombination with other mobile genetic elements. is a clinically important pathogen representing a serious threat to human health. Our hypothesis is that genetic differences between strains caused by the presence of integrated prophages could explain the apparent differences observed in the virulence of different strains. In this study, we provide a full characterization of phiCD211, also known as phiCDIF1296T, the largest phage known to infect so far. Screening 2,584 genomes revealed the presence of highly similar phiCD211-like phages in 5% of the strains analyzed, showing their high prevalence. Multiple-genome comparisons suggest that evolution of the phiCD211-like phage community is dynamic, and some members have acquired genes that could influence bacterial biology and fitness. Our study further supports the relevance of studying phages in to better understand the epidemiology of this clinically important human pathogen.

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“…Several genetic markers are currently available for discrimination of C. difficile strains, but they vary in efficiency. Plasmid analysis has been first developed but this typing method is not suitable to all C. difficile strains because only 40% have plasmidic materials [12]. Phage typing is a long and time consuming technique and it can be performed only in reference laboratories [12].…”

Section: Discussionmentioning

confidence: 99%

“…Plasmid analysis has been first developed but this typing method is not suitable to all C. difficile strains because only 40% have plasmidic materials [12]. Phage typing is a long and time consuming technique and it can be performed only in reference laboratories [12]. The digestion of genomic DNA with restriction enzymes produces a large number of bands making the interpretation of DNA patterns difficult [13].…”

Section: Discussionmentioning

confidence: 99%

Genomic fingerprinting ofClostridium difficileisolates by using a random amplified polymorphic DNA (RAPD) assay

Barbut¹,

Mario²,

Delmée³

et al. 1993

FEMS Microbiology Letters

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This study describes the use of a new and easy method called random amplified polymorphic DNA (RAPD) assay to distinguish strains of C. difficile. We used two single short primers (AP4 and AP5) with arbitrary nucleotide sequences in a polymerase chain reaction to amplify genomic DNA. The profiles observed after electrophoretic separation were able to distinguish 20 reference C. difficile strains previously serotyped by Delmée's method. The fingerprints of 11 epidemiologically unrelated C. difficile strains clearly yielded a DNA polymorphism between all the strains. Latterly, RAPD profiles of 11 C. difficile strains isolated from 2 independent suspected outbreaks showed, in each case, a predominant banding pattern corresponding to an epidemic strain. These results suggest that RAPD assay could be a valuable tool for epidemiological studies.

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Development and application of a multiple typing system for Clostridium difficile

Cited by 32 publications

References 31 publications

Comparison of restriction endonuclease analysis, ribotyping, and pulsed-field gel electrophoresis for molecular differentiation of Clostridium difficile strains

Comparison of restriction endonuclease analysis, ribotyping, and pulsed-field gel electrophoresis for molecular differentiation of Clostridium difficile strains

High Prevalence and Genetic Diversity of Large phiCD211 (phiCDIF1296T)-Like Prophages in Clostridioides difficile

Genomic fingerprinting ofClostridium difficileisolates by using a random amplified polymorphic DNA (RAPD) assay

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