1887

Abstract

infections can be effectively treated with clarithromycin, a macrolide, in combination with other antibiotics, such as amoxicillin, tetracycline or metronidazole. The failure of eradication is mainly associated with macrolide-resistant strains. Three point mutations (A2142G/C, A2143G, T2182C) in the peptidyltransferase region of domain V of the 23S rRNA have been described as being associated with clarithromycin resistance. Therefore, the determination of clarithromycin resistance by pyrosequencing was evaluated. from 81 gastric biopsies was cultured and clarithromycin resistance was determined by Etest, as well as by pyrosequencing technology (PSQ 96 system; Biotage). The respective mutations were set in relation to the MIC measured in μg ml by Etest. In this study, point mutations in positions 2142 and 2143 were associated with clarithromycin resistance. Mutations in position 2182 did not contribute to clarithromycin resistance. In addition, from 22 out of the 81 biopsies, clarithromycin resistance was determined directly without culturing to save additional time. Identical results were obtained as compared to resistance testing with pure strains. All results obtained by pyrosequencing were evaluated by Sanger sequencing. The data show that pyrosequencing to detect point mutation is a fast and reliable method for determining clarithromycin resistance in , and provides the same results as the Etest.

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2007-10-01
2019-10-21
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References

  1. Alarcon, T., Domingo, D., Prieto, N. & Lopez-Brea, M. ( 2000; ). PCR using 3′-mismatched primers to detect A2142C mutation in 23S rRNA conferring resistance to clarithromycin in Helicobacter pylori clinical isolates. J Clin Microbiol 38, 923–925.
    [Google Scholar]
  2. Debets-Ossenkopp, Y. J., Brinkman, A. B., Kuipers, E. J., Vandenbroucke-Grauls, C. M. J. E. & Kusters, J. G. ( 1998; ). Explaining the bias in the 23S rRNA gene mutations associated with clarithromycin resistance in clinical isolates of Helicobacter pylori. Antimicrob Agents Chemother 42, 2749–2751.
    [Google Scholar]
  3. Debets-Ossenkopp, Y. J., Herscheid, A. J., Pot, R. G., Kuipers, E. J., Kusters, J. G. & Vandenbroucke-Grauls, C. M. ( 1999; ). Prevalence of Helicobacter pylori resistance to metronidazole, clarithromycin, amoxycillin, tetracycline and trovafloxacin in The Netherlands. J Antimicrob Chemother 43, 511–515.[CrossRef]
    [Google Scholar]
  4. Fontana, C., Favaro, M., Minelli, S., Criscuolo, A. A., Pietroiusti, A., Galante, A. & Favalli, C. ( 2002; ). New site of modification of 23S rRNA associated with clarithromycin resistance of Helicobacter pylori clinical isolates. Antimicrob Agents Chemother 46, 3765–3769.[CrossRef]
    [Google Scholar]
  5. Fontana, C., Favaro, M., Pietroiusti, A., Pistoia, E. S., Galante, A. & Favalli, C. ( 2003; ). Detection of clarithromycin-resistant Helicobacter pylori in stool samples. J Clin Microbiol 41, 3636–3640.[CrossRef]
    [Google Scholar]
  6. Haanpera, M., Huovinen, P. & Jalava, J. ( 2005; ). Detection and quantification of macrolide resistance mutations at positions 2058 and 2059 of the 23S rRNA gene by pyrosequencing. Antimicrob Agents Chemother 49, 457–460.[CrossRef]
    [Google Scholar]
  7. Heep, M., Kist, M., Strobel, S., Beck, D. & Lehn, N. ( 2000; ). Secondary resistance among 554 isolates of Helicobacter pylori after failure of therapy. Eur J Clin Microbiol Infect Dis 19, 538–541.[CrossRef]
    [Google Scholar]
  8. Hjalmarsson, S., Alderborn, A., Fock, C., Muldin, I., Kling, H., Uhlén, M. & Engstrand, L. ( 2004; ). Rapid combined characterization of microorganism and host genotypes using a single technology. Helicobacter 9, 138–145.[CrossRef]
    [Google Scholar]
  9. Hultén, K., Gibreel, A., Sköld, O. & Engstrand, L. ( 1997; ). Macrolide resistance in Helicobacter pylori: mechanism and stability in strains from clarithromycin-treated patients. Antimicrob Agents Chemother 41, 2550–2553.
    [Google Scholar]
  10. Jiang, Q., Hiratsuka, K. & Taylor, D. E. ( 1996; ). Variability of gene order in different Helicobacter pylori strains contributes to genome diversity. Mol Microbiol 20, 833–842.[CrossRef]
    [Google Scholar]
  11. Khan, R., Nahar, S., Sultana, J., Ahmad, M. M. & Rahman, M. ( 2004; ). T2182C mutation in 23S rRNA is associated with clarithromycin resistance in Helicobacter pylori isolates obtained in Bangladesh. Antimicrob Agents Chemother 48, 3567–3569.[CrossRef]
    [Google Scholar]
  12. Kim, K. S., Kang, J. O., Eun, C. S., Han, D. S. & Choi, T. Y. ( 2002; ). Mutations in the 23S rRNA gene of Helicobacter pylori associated with clarithromycin resistance. J Korean Med Sci 17, 599–603.[CrossRef]
    [Google Scholar]
  13. Kist, M. ( 1991; ). Diagnostische Verfahrensrichtlinien der DGHM: Isolierung und Identifizierung von Bakterien der Gattungen Campylobacter und Helicobacter. Zentralbl Bakteriol 276, 124–139.[CrossRef]
    [Google Scholar]
  14. Kobayashi, I., Saika, T., Muraoka, H., Murakami, K. & Fujioka, T. ( 2006; ). Helicobacter pylori isolated from patients who later failed H. pylori eradication triple therapy readily develop resistance to clarithromycin. J Med Microbiol 55, 737–740.[CrossRef]
    [Google Scholar]
  15. Marais, A., Monteiro, L., Occhialini, A., Pina, M., Lamouliatte, H. & Megraud, F. ( 1999; ). Direct detection of Helicobacter pylori resistance to macrolides by a polymerase chain reaction DNA enzyme immunoassay in gastric biopsy specimens. Gut 44, 463–467.[CrossRef]
    [Google Scholar]
  16. Occhialini, A., Urdaci, M., Doucet-Populaire, F., Bebear, C. M., Lamouliatte, H. & Megraud, F. ( 1997; ). Macrolide resistance in Helicobacter pylori: rapid detection of point mutations and assays of macrolide binding to ribosomes. Antimicrob Agents Chemother 41, 2724–2728.
    [Google Scholar]
  17. Owen, R. J. ( 2002; ). Molecular testing for antibiotic resistance in Helicobacter pylori. Gut 50, 285–289.[CrossRef]
    [Google Scholar]
  18. Pina, M., Occhialini, A., Monteiro, L., Doermann, H. P. & Megraud, F. ( 1998; ). Detection of point mutations associated with resistance of Helicobacter pylori to clarithromycin by hybridization in liquid phase. J Clin Microbiol 36, 3285–3290.
    [Google Scholar]
  19. Ronaghi, M. ( 2000; ). Improved performance of pyrosequencing using single-stranded DNA-binding protein. Anal Biochem 286, 282–288.[CrossRef]
    [Google Scholar]
  20. Ronaghi, M. ( 2001; ). Pyrosequencing sheds light on DNA sequencing. Genome Res 11, 3–11.[CrossRef]
    [Google Scholar]
  21. Ronaghi, M., Karamohamed, S., Pettersson, B., Uhlen, M. & Nyren, P. ( 1996; ). Real-time DNA sequencing using detection of pyrophosphate release. Anal Biochem 242, 84–89.[CrossRef]
    [Google Scholar]
  22. Ronaghi, M., Uhlen, M. & Nyren, P. ( 1998; ). A sequencing method based on real-time pyrophosphate. Science 281, 363–365.[CrossRef]
    [Google Scholar]
  23. Ronaghi, M., Nygren, M., Lundeberg, J. & Nyren, P. ( 1999; ). Analyses of secondary structures in DNA by pyrosequencing. Anal Biochem 267, 65–71.[CrossRef]
    [Google Scholar]
  24. Sinclair, A., Arnold, C. & Woodford, N. ( 2003; ). Rapid detection and estimation by pyrosequencing of 23S rRNA genes with a single nucleotide polymorphism conferring linezolid resistance in enterococci. Antimicrob Agents Chemother 47, 3620–3622.[CrossRef]
    [Google Scholar]
  25. Strobel, S., Bereswill, S., Balig, P., Allgaier, P., Sonntag, H. G. & Kist, M. ( 1998; ). Identification and analysis of a new vacA genotype variant of Helicobacter pylori in different patient groups in Germany. J Clin Microbiol 36, 1285–1289.
    [Google Scholar]
  26. Szczebara, F., Dhaenens, L., Vincent, P. & Husson, M. O. ( 1997; ). Evaluation of rapid molecular methods for detection of clarithromycin resistance in Helicobacter pylori. Eur J Clin Microbiol Infect Dis 16, 162–164.[CrossRef]
    [Google Scholar]
  27. Taylor, D. E., Ge, Z., Purych, D., Lo, T. & Hiratsuka, K. ( 1997; ). Cloning and sequence analysis of two copies of a 23S rRNA gene from Helicobacter pylori and association of clarithromycin resistance with 23S rRNA mutations. Antimicrob Agents Chemother 41, 2621–2628.
    [Google Scholar]
  28. Toracchio, S., Cellini, L., Di Campli, E., Cappello, G., Malatesta, M. G., Ferri, A., Ciccaglione, A. F., Grossi, L. & Mario, L. ( 2000; ). Role of antimicrobial susceptibility testing on efficacy of triple therapy in Helicobacter pylori eradication. Aliment Pharmacol Ther 14, 1639–1643.[CrossRef]
    [Google Scholar]
  29. Treiber, G., Kist, M., Klotz, U., Peitz, U. & Malfertheiner, P. ( 2005; ). Therapie der Helicobacter-pylori-infektion: klinische, mikrobiologische und pharmakologische Aspekte. Dtsch Arztebl 26, 1883–1888.
    [Google Scholar]
  30. van Doorn, L. J., Glupczynski, Y., Kusters, J. G., Megraud, F., Midolo, P., Maggi-Scola, N., Queiroz, D. M., Nouhan, N., Stet, E. & Quint, W. G. ( 2001; ). Accurate prediction of macrolide resistance in Helicobacter pylori by a PCR line probe assay for detection of mutations in the 23S rRNA gene: multicenter validation study. Antimicrob Agents Chemother 45, 1500–1504.[CrossRef]
    [Google Scholar]
  31. Versalovic, J., Shortridge, D., Kibler, K., Griffy, M. V., Beyer, J., Flamm, R. K., Tanaky, S. K., Graham, D. Y. & Go, M. F. ( 1996; ). Mutations in 23S rRNA are associated with clarithromycin resistance in Helicobacter pylori. Antimicrob Agents Chemother 40, 477–480.
    [Google Scholar]
  32. Xia, H. X., Buckley, M., Keane, C. T. & O'Morain, C. A. ( 1996; ). Clarithromycin resistance in Helicobacter pylori: prevalence in untreated dyspeptic patients and stability in vitro. J Antimicrob Chemother 37, 473–481.[CrossRef]
    [Google Scholar]
  33. Yahav, J., Samra, Z., Niv, Y., Evans, C. T., Passaro, D. J., Dinari, G. & Shmuely, H. ( 2006; ). Susceptibility-guided vs. empiric retreatment of Helicobacter pylori infection after treatment failure. Dig Dis Sci 51, 2316–2321.[CrossRef]
    [Google Scholar]
  34. Zhao, J. R., Bai, Y. J., Zhang, Q. H., Wang, Y., Luo, M. & Yan, X. J. ( 2005; ). Pyrosequencing-based approach for rapid detection of rifampin-resistant Mycobacterium tuberculosis. Diagn Microbiol Infect Dis 51, 135–137.[CrossRef]
    [Google Scholar]
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