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Antimicrobial Agents and Chemotherapy, May 2001, p. 1500-1504, Vol. 45, No. 5
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.5.1500-1504.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

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

Leen-Jan van Doorn,1,* Youri Glupczynski,2 Johannes G. Kusters,3 Francis Mégraud,4 Peter Midolo,5 Nadia Maggi-Solcà,6 Dulciene M. M. Queiroz,7 Nathalie Nouhan,1 Els Stet,1 and Wim G. V. Quint1

Delft Diagnostic Laboratory, Delft, The Netherlands1; Cliniques Universitaire UCL de Mont-Godinne, Yvoir, Belgium2; Department of Medical Microbiology, Free University, Amsterdam, The Netherlands3; Laboratoire de Bactériologie, Hôpital Pellegrin, Bordeaux, France4; Monash Medical Center, Clayton, Australia5; Istituto Cantonale Batteriosierologico, Lugano, Switzerland6; and Laboratory for Research in Bacteriology, Faculty of Medicine/UFMG, Belo Horizonte, Brazil7

Received 18 September 2000/Returned for modification 4 January 2001/Accepted 9 February 2001

Helicobacter pylori strains from 299 patients were tested in six laboratories in different countries. Macrolide susceptibility of the strains was determined by agar dilution (17.4%) or the epsilometer test (82.6%). Mutations in the 23S ribosomal DNA (rDNA) that are associated with macrolide resistance were analyzed by PCR and reverse hybridization (PCR-line probe assay [LiPA]). This method identifies A2115G, G2141A, A2142G, A2142C, A2142T, A2143G, and A2143C mutations in the 23S rDNA. vacA s-region (s1a, s1b, s1c, and s2) and m-region (m1, m2a, and m2b) genotypes and cagA status were also determined using another PCR-LiPA system. Of the 299 strains investigated by MIC testing, 130 (43.5%) were resistant and 169 (56.5%) were susceptible to clarithromycin. Of the 130 resistant strains, 127 (97.7%) contained 23S rDNA mutations, whereas 167 (98.8%) of the 169 susceptible strains contained wild-type sequences. The predominant mutations were A2143G (45.2%) and A2142G (33.3%). Twenty-eight (19.8%) strains contained multiple 23S rDNA mutations. Only five resistant strains contained the A2142C mutation (three of these in combination with the A2142G mutation), and the A2115G, G2141A, A2142T, and A2143C mutations were not found. MICs of clarithromycin for the A2142G mutant strains were significantly higher than MICs for the A2143G strains. Although there was no significant association between 23S rDNA mutations and the vacA and cagA status, clarithromycin-susceptible strains more often contained mixed vacA genotypes, indicating the presence of multiple H. pylori strains. In conclusion, our data confirmed the very strong association between 23S rDNA mutations and macrolide resistance and showed that the PCR-LiPA permits accurate and reliable diagnosis of macrolide resistance in H. pylori.

* Corresponding author. Mailing address: Delft Diagnostic Laboratory, R. de Graafweg 7, 2625 AD, Delft, The Netherlands. Phone: 31-15-2604581. Fax: 31-15-2604550. E-mail: L.J.van.Doorn{at}ddl.nl.

Antimicrobial Agents and Chemotherapy, May 2001, p. 1500-1504, Vol. 45, No. 5
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.5.1500-1504.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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