Canadian Journal of Gastroenterology and Hepatology

Canadian Journal of Gastroenterology and Hepatology / 2009 / Article

Original Article | Open Access

Volume 23 |Article ID 695349 | 4 pages | https://doi.org/10.1155/2009/695349

Pyrosequencing Assay to Rapidly Detect Clarithromycin Resistance Mutations in Canadian Helicobacter pylori Isolates

Received31 Jul 2008
Accepted02 Dec 2008

Abstract

BACKGROUND: Mutations at positions 2142 or 2143 in the two-copy 23S ribosomal RNA gene of Helicobacter pylori are highly predictive of in vitro clarithromycin resistance and failure of clarithromycin-containing treatment regimens.OBJECTIVE: To design an assay to rapidly detect these mutations using rapid polymerase chain reaction and pyrosequencing, a novel method of ‘sequencing by synthesis’, and to test this assay with a collection of Canadian H pylori isolates.METHODS: Forty-two H pylori isolates (24 clarithromycin-resistant, 18 clarithromycin-susceptible) were studied. A target region in the 23S gene was rapidly amplified and sequenced by pyrosequencing.RESULTS: Mutations at one of the two positions studied were present in 20 of the 24 (83%) clarithromycin-resistant isolates; 13 had double-copy A2143G mutations, four had double-copy A2142G mutations and three had single-copy A2143G mutations. There were no mutations in 17 of the 18 (94%) susceptible isolates. A single-copy A2142G mutation was detected in one susceptible isolate.CONCLUSIONS: The pyrosequencing assay developed was able to detect and differentiate mutations at positions 2142 and 2143 in either one or both copies of the H pylori 23S ribosdomal RNA gene. Further study is needed to determine whether this pyrosequencing assay can be used to determine H pylori susceptibility to clarithromycin from clinical specimens such as stools or gastric biopsies.

Copyright © 2009 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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