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Letters to the Editor Genotyping of Clostridium difficile Isolates



With regard to the recent report (2) on typing of Clostridium difficile by arbitrarily primed PCR (AP-PCR), we would like to make readers aware of another typing approach (1). This approach uses PCR to amplify the 16S-23S rRNA gene (rDNA) spacer region, which is situated in the evolutionarily stable rRNA operon (5). It was demonstrated that in C. difficile there were 16 variable-length 16S-23S rDNA spacer alleles: different strains contained different combinations of variable-length alleles (1). By AP-PCR, 41 isolates were separated into nine groups, with 66 % falling into one group (2). In a separate study, AP-PCR was used to separate 20 isolates into four groups (3). By amplifying the 16S-23S rDNA spacer region (1), 24 isolates were divided into 14 ribotypes. Although the total number of genotypes is not known, the 16S-23S rDNA spacer typing is more discriminatory than AP-PCR typing. The 16S-23S spacer region typing is based on PCR, and so the laboratory time taken to type organisms is comparable to that taken with AP-PCR: about 1.5 working days from extrac-tion to gel reading. AP-PCR is based on the partial homology of short primers (5 to 10 nucleotides) to genomic DNA, resulting in primers randomly annealing (4): the method produces results which vary from run to run and from operator to operator and are highly dependent on PCR conditions. On the other hand, 16S-23S spacer typing is based on the presence or absence of long regions ofDNA (150 to 500 bp) in an operon that is stable in evolutionary terms (1, 5). Taking this and the improved discriminatory power together, 16S-23S rDNA spacer region typing has advantages over AP-PCR

Year: 2016
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