Survey of the extrachromosomal gene pool of Clostridium difficile

Muldrow, Lycurgus L.; Archibold, Errol R.; Nunez-Montiel, O; Sheehy, Ronald J.

doi:10.1128/jcm.16.4.637-640.1982

Cited by 37 publications

(12 citation statements)

References 28 publications

(20 reference statements)

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“…The standard cytotoxicity and antibody neutralization assays were performed to test toxin activity using chinese hamster ovary (CHO) cells as described previously [8,9]. Toxic activity was expressed as the greatest dilution that caused 50% cell death.…”

Section: Cytotoxic Assaysmentioning

confidence: 99%

Molecular screening of Clostridium difficile toxins A and B genetic determinants and identification of mutant strains

McMillin

1991

FEMS Microbiology Letters

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Three separate sets of polymerase chain reaction primers were designed to specifically detect the presence of a toxin A gene fragment, a toxin B gene fragment, and the entire toxin B gene. In addition toxin gene fragments that were amplified from well characterized toxic strains were tagged fluorescently and used as hybridization probes to screen C. difficile strains. A survey of 37 toxic strains and 10 non-toxic strains demonstrated that toxic strains normally contain the genetic composition for toxin A and toxin B simultaneously; whereas, non-toxic strains typically did not contain detectable toxin determinants. The only exception found was strain 39, which had the genetic composition for toxins A and B, but was not cytotoxic under the conditions tested.

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Section: Cytotoxic Assaysmentioning

confidence: 99%

Molecular screening of Clostridium difficile toxins A and B genetic determinants and identification of mutant strains

McMillin

1991

FEMS Microbiology Letters

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“…This bacterium is considered to be the main agent of nosocomially acquired diarrhea among adults [2]. Many various typing methods have been developed to investigate nosocomial outbreaks of C. di¤cile, some based on phenotype (lysotyping [3], serogroup-ing [4], SDS-PAGE [5], immunoblotting [6]) and others on genotype (plasmids [7], REA [8], ribotyping [9], PFGE [10], RAPD [11]). Recently, Kostman et al developed a method of PCR-ribotyping that was shown to be reproducible, easy to perform and cost-e¡ective [12].…”

Section: Introductionmentioning

confidence: 99%

Development of a new PCR-ribotyping method for Clostridium difficile based on ribosomal RNA gene sequencing

Bidet

1999

FEMS Microbiology Letters

101

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PCR-ribotyping, a typing method based on polymorphism in the 16S-23S intergenic spacer region, has been recently used to investigate outbreaks due to Clostridium difficile. However, this method generates bands of high and close molecular masses which are difficult to separate on agarose gel electrophoresis. To improve reading of banding patterns of PCR-ribotyping applied to C. difficile, a partial sequencing of the rRNA genes (16S and 23S) and intergenic spacer region has been performed, then a new set of primers located closer to the intergenic spacer region has been defined. The new PCR gave reproducible patterns of bands easy to separate on agarose gel electrophoresis. Each of the 10 serogroups and 11 subgroups of serogroup A produced a different pattern. This typing method has evidenced major qualities such as easiness, rapidity and reproducibility. However, its discriminatory power has to be evaluated to validate its importance as a typing tool for C. difficile. z

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“…When isolates from 35 geographically different sources were tested, 24 (66%) had plasmids, i.e., were typeable. Muldrow et al (19) found that only 18% of the 82 strains they analyzed carried plasmids. The plasmids ranged from 2.7 to 60 MDa in size.…”

Section: Discussionmentioning

confidence: 99%

Development and application of a multiple typing system for Clostridium difficile

Mahony

Clow

Atkinson

et al. 1991

Appl Environ Microbiol

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A combination of bacteriocin, bacteriophage, and plasmid typing techniques was used to differentiate strains of Clostridium difficile. A typing set of 20 bacteriocin-producing strains was established after 400 isolates of C. difficile were screened for the ability to produce bacteriocin. These strains were used to type a collection of 114 isolates of C. difficile. Forty-six (40%) of the 114 isolates were typeable, and 31 typing patterns were distinguishable. Plasmid typing of the same 114 isolates of C. difficile showed that 67 (59%) of the isolates carried up to four plasmids ranging from 7 to 60 kb in size, although most strains contained only one or two plasmids. Twenty different plasmid typing patterns were observed among the isolates. A combination of bacteriocin and plasmid typing provided 77% typeability. Fifteen (13%) of the 114 strains were typeable with five bacteriophages isolated in our laboratory, but the increase in typeability of strains over that obtainable by plasmid and bacteriocin typing was only 1.8%. Isolates that were nontypeable by bacteriocins, plasmids, or phages could be divided into two groups on the basis of positive or negative cytotoxin production. This further division of strains would increase the typeability potential by 7%; i.e., the ability to differentiate strains would rise from 77 to 84%, or perhaps 86%, if phage typing were included. We conclude that more than one of the techniques reported in this paper must be used to achieve an acceptable level of typeability of this species.

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Survey of the extrachromosomal gene pool of Clostridium difficile

Cited by 37 publications

References 28 publications

Molecular screening of Clostridium difficile toxins A and B genetic determinants and identification of mutant strains

Molecular screening of Clostridium difficile toxins A and B genetic determinants and identification of mutant strains

Development of a new PCR-ribotyping method for Clostridium difficile based on ribosomal RNA gene sequencing

Development and application of a multiple typing system for Clostridium difficile

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