Toxinotyping and PCR ribotyping are two methods that have been used to type Clostridium difficile isolates. Toxinotyping is based on PCR-RFLP analysis of a 19 kb region encompassing the C. difficile pathogenicity locus. PCR ribotyping is based on comparison of patterns of PCR products of the 16S–23S rRNA intergenic spacer region. Representative strains (101) from a C. difficile PCR ribotype library and 22 strains from previously described toxinotypes were analysed to compare ribotyping with toxinotyping. Within this panel of strains all 11 toxinotypes (0–X) described previously and an additional 5 novel toxinotypes (XI–XV) were observed. PCR ribotyping and toxinotyping correlated well and usually all strains within a given ribotype had similar changes in toxin genes. The new toxinotype XI comprises strains that did not express toxins TcdA or TcdB at detectable levels, but contained part of the tcdA gene. Strains of toxinotype XII exhibit changes only in the 5′ end of the tcdB gene. Toxinotype XIV is composed of strains that have a large insertion at the beginning of the tcdA gene. A total of 25 of the 89 tested PCR ribotypes of C. difficile contained variant strains. It was estimated that they represent 7·7% of the total number of strains in the Anaerobe Reference Unit collection.
BarbutF. N., MarioJ., FrottierJ., PetitJ. C.1993; Use of arbitrary primer PCR for investigating an outbreak of Clostridium difficile-associated diarrhoea in AIDS patients. Eur J Clin Microbiol Infect Dis 12:724–795
BraunV., HundsbergerT., LeukelP., SauerbornM., von Eichel-StreiberC.1996; Definition of the single integration site of the pathogenicity locus in Clostridium difficile. Gene 181:29–38[CrossRef]
BraunV., MehligM., MoosM., RupnikM., KaltB., MahonyD. E., von Eichel-StreiberC.2000; A chimeric ribozyme in Clostridium difficile combines features of group I introns and insertion elements. Mol Microbiol 36:1447–1459
BrazierJ. S., MulliganM. E., Delm
é
eM., TabaqchaliS.1997; Preliminary findings of the international typing study on Clostridium difficile. International Clostridium Difficile Study Group. Clin Infect Dis 25:Suppl 2S199–S201[CrossRef]
CohenS. H., TangY. J., HansenB., SilvaJ.Jr1998; Isolation of a toxin B-deficient mutant strain of Clostridium difficile in a case of recurrent C. difficile-associated diarrhea. Clin Infect Dis 26:410–412[CrossRef]
DelméeM., LarocheY., AvesaniV., CornelisG.1986; Comparison of serogrouping and polyacrylamide gel electrophoresis for typing of Clostridium difficile. J Clin Microbiol 24:991–994
DepitreC., DelméeM., AvesaniV., L’HaridonR., RoelsA., PopoffM., CorthierG.1993; Serogroup F strains of Clostridium difficile produce toxin B but not toxin A. J Med Microbiol 38:434–441[CrossRef]
van DijckP., AvesaniV., DelméeM.1996; Genotyping of outbreak-related and sporadic isolates of Clostridium difficile belonging to serogroup C. J Clin Microbiol 34:3049–3055
von Eichel-StreiberC., SauerbornM.1990; Clostridium difficile toxin A carries a C-terminal repetitive structure homologous to the carbohydrate binding region of streptococcal glycosyltransferases. Gene 96:107–113[CrossRef]
von Eichel-StreiberC., BoquetP., SauerbornM., ThelestamM.1996; Large clostridial cytotoxins – a family of glycosyltransferases modifying small GTP-binding proteins. Trends Microbiol 4:375–382[CrossRef]
von Eichel-StreiberC., Zec-PirnatI., GrabnarM., RupnikM.1999; A nonsense mutation abrogates production of functional enterotoxin A in Clostridium difficile toxinotype VIII strains of serogroups F and X. FEMS Microbiol Lett 178:163–168[CrossRef]
HutsonR. A., ThompsonD. E., CollinsM. D.1993; Genetic interrelationships of saccharolytic Clostridium botulinum types B, E and F and related clostridia as revealed by small-subunit rRNA gene sequences. FEMS Microbiol Lett 108:103–110[CrossRef]
LyerlyD. M., BarrosoL. A., WilkinsT. D., DepitreC., CorthierG.1992; Characterization of a Toxin A-negative, Toxin B-positive strain of Clostridium difficile. Infect Immun 60:4633–4639
MathisJ. N., PilkintonL., McMillinD. E.1999; Detection and transcription of toxin DNA in a nontoxigenic strain of Clostridium difficile. Curr Microbiol 38:324–328[CrossRef]
MulliganM. E., PetersonL. R., KwokR. Y. Y., ClabotsC. R., GerdingD. N.1988; Immunoblots and plasmid fingerprints compared with serotyping and polyacrylamide gel electrophoresis for typing Clostridium difficile. J Clin Microbiol 26:41–46
O’NeillG. L., OgunsolaF. T., BrazierJ. S., DuerdenB. I.1996; Modification of a PCR rIbotyping method for application as a routine typing scheme for Clostridium difficile. Anaerobe 2:205–209[CrossRef]
RupnikM., BraunV., SoehnF., JancM., HofstetterM., Laufenberg-FeldmanR., von Eichel-StreiberC.1997; Characterization of polymorphisms in the toxin A and B genes of Clostridium difficile. FEMS Microbiol Lett 148:197–202[CrossRef]
RupnikM., AvesaniV., JancM., von Eichel-StreiberC., DelméeM.1998; A novel toxinotyping scheme and correlation of toxinotypes with serogroups of Clostridium difficile isolates. J Clin Microbiol 36:2240–2247
SoehnF., Wagenknecht-WiesnerA., LeukelP., KohlM., WeidmanM., von Eichel-StreiberC., BraunV.1998; Genetic rearrangements in the pathogenicity locus of Clostridium difficile strain 8864 – implications for transcription, expression and enzymatic activity of toxins A and B. Mol Gen Genet 258:222–232[CrossRef]
StubbsS. L. J., BrazierJ. S., O’NeillG. L., DuerdenB. I.1999; PCR targeted to the 16S–23S rRNA gene intergenic spacer region of Clostridium difficile and construction of a library consisting of 116 different PCR ribotypes. J Clin Microbiol 37:461–463