1Microbiology Laboratories, 2Helicobacter pylori Research Institute and 3Department of Gastroenterology, Rabin Medical Center, Beilinson Campus, Petah Tiqva; 4Institute of Pediatric Gastroenterology and Nutrition, Schneider Childrens Medical Center of Israel, Petah Tiqva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel; 5Division of Epidemiology and Biostatistics, University of Illinois, Chicago, IL, USA
Received 13 November 2001; returned 21 January 2002; revised 18 February 2002; accepted 27 February 2002.
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Abstract |
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Introduction |
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The aim of this study was to assess the resistance of H. pylori isolates from untreated and treated patients in Israel to five antimicrobial agents. Cefixime was included because previous investigations have found it to possess anti-H. pylori activity both in vivo and in vitro.6
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Materials and methods |
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Between January 2000 and June 2001, 138 clinical isolates of H. pylori were isolated from antral biopsy specimens of 138 dyspeptic adults (71 females, 67 males; median age 55 years), one isolate from each patient, at the Rabin Medical Center, a major tertiary hospital in central Israel. Twenty-eight patients had been treated previously for H. pylori infection with a PPI and two or more antimicrobial agents: 15 with amoxicillin, clarithromycin and metronidazole; seven with clarithromycin and metronidazole, three with amoxicillin and clarithromycin; two with clarithromycin, metronidazole and tetracycline; and one with amoxicillin and metronidazole.
The specimens were inoculated directly on to Columbia blood agar (Difco, Detroit, MI, USA) supplemented with yeast extract (5 g/L), laked lysed horse blood (7%), vancomycin (3 mg/L), colistin sulphate (7.5 mg/L), nystatin (12 500 IU/L) and co-trimoxazole (5 mg/L). Cultures were incubated for 72 h at 37°C under microaerophilic conditions. H. pylori isolates were identified by colony morphology, characteristic spiral morphology on Gram staining, and positive findings for catalase, urease and oxidase tests.
MIC determination
The MICs of amoxicillin, clarithromycin, metronidazole, tetracycline and cefixime were determined by Etest (AB Biodisk, Solna, Sweden). Suspensions to a density of a 3 McFarland standard were prepared in Columbia broth (Difco) from 72 h blood agar subculture and spread (100 µL) on Petri plates (90 mm diameter) containing MuellerHinton agar (BBL, Becton Dickinson Microbiology Systems, Cockeysville, MD, USA) supplemented with yeast extract (5 g/L), horse serum (12%) and nicotinamide adenine dinucleotide (25 mg/L). Etest strips were placed on the plates as soon as the inocula were absorbed into the agar. The plates were incubated at 37°C under microaerophilic conditions. MIC values were determined after 72 h incubation. Resistance was defined as follows: amoxicillin, MIC 1.5 mg/L; tetracycline
4 mg/L; clarithromycin
2 mg/L; metronidazole
8 mg/L7 and cefixime
2 mg/L. H. pylori strain ATCC 43526 was used for quality control of the selective medium and ATCC 43504 for the quality control of the susceptibility tests.
Prevalence of resistance was calculated between treated and untreated patients for each antibiotic with the MantelHaenszel 2 test or Fishers exact test. Cornfield or exact 95% confidence intervals (CIs) were calculated as appropriate.
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Results |
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Discussion |
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The clinical significance of the resistance level of H. pylori on in vitro studies has long been disputed. Nevertheless, recent meta-analyses have provided conclusive evidence that H. pylori resistance to metronidazole or clarithromycin reduces the efficacy of regimens containing these agents.8 As antimicrobial resistance is the most important determinant after patient compliance of outcome of treatment,9 and metronidazole and clarithromycin are used in almost all regimens, the increasing resistance of H. pylori to these agents is of clinical importance. In our study, all clarithromycin- or metronidazole-resistant isolates from the treated group were recovered from patients treated previously with clarithromycin or metronidazole, respectively.
No resistance to tetracycline was found, and the sole strain that was resistant to amoxicillin was recovered from an untreated patient (MIC = 1.5 mg/L). Tetracycline possessed the most significant anti-H. pylori activity in vitro, and it may be used as an alternative agent in the presence of clarithromycin and/or metronidazole resistance in adult patients.
Cefixime was found to be highly active against clarithromycin-resistant strains of H. pylori. It had similar activity to tetracycline against both clarithromycin-sensitive and clarithromycin-resistant H. pylori isolates from untreated and treated patients.
Dual resistance to metronidazole and clarithromycin was common (28%) in isolates from treated patients. Previous studies have shown that for patients infected by clarithromycin-resistant strains, the use of treatment regimens containing both metronidazole and clarithromycin dramatically decreases the H. pylori eradication rate.10 Thus, in countries where metronidazole resistance is common, physicians might well refrain from using a combination of metronidazole and clarithromycin in order to prevent the emergence of more resistant strains.
In conclusion, the high prevalence of both single and dual metronidazole and clarithromycin resistance in isolates recovered from treated patients warrants special attention. Tetracycline and cefixime are potential alternative agents for the treatment of clarithromycin- and metronidazole-resistant H. pylori infection. Further clinical studies are needed to confirm these data in vivo.
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Acknowledgements |
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Footnotes |
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References |
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