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2014 | 61 | 2 | 311-315
Article title

PCR-RFLP detection of point mutations A2143G and A2142G in 23S rRNA gene conferring resistance to clarithromycin in Helicobacter pylori strains

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Background. The occurrence of clarithromycin resistance among Helicobacter pylori strains is a major cause of the treatment failure. Resistance to this drug is conferred by point mutations in 23S rRNA gene and the most prevalent mutations are A2143G and A2142G. The aim of the study was to evaluate the occurrence of A2143G and A2142G mutations in a group of H. pylori strains resistant to clarithromycin. Materials and Methods. The study included 21 clarithromycin-resistant H. pylori strains collected between 2006 and 2009 in southern Poland. Resistance to clarithromycin was quantitatively tested with the E-test to determine the minimal inhibitory concentration (MIC value). The point mutations of H. pylori isolates were detected by PCR followed by RFLP analysis. Results. The MIC values for clarithromycin for the analyzed strains ranged from 1.5 mg/L to 64 mg/L. Nine H. pylori strains exhibited A2143G mutation and A2142G mutation was found in 9 isolates as well. The results of RFLP analysis of 3 clarithromycin-resistant strains were negative for both mutations. The average MIC values for A2143G and A2142G mutants were 6 and 30 mg/L, respectively. Conclusions. Frequencies of A2143G and A2142G mutations were the same in all isolates tested. Strains with A2143G mutation exhibited lower MIC values than A2142G mutants. Application of PCR-RFLP method for detection of clarithromycin resistance allows for better and more efficient management of H. pylori infections.
Physical description
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Falck Medycyna Outpatient Clinic of Gastroenterology, Kraków, Poland
  • Department of Pharmaceutical Microbiology, Jagiellonian University Medical College, Kraków, Poland
  • Agudo S, Alarcón T, Urruzuno P, Martínez MJ, López-Brea M (2010) Detection of Helicobacter pylori and clarithromycin resistance in gastric biopsies of pediatric patients by using a commercially available real-time polymerase chain reaction after NucliSens semiautomated DNA extraction. Diagn Microbiol Infect Dis 67: 213-219.
  • Agudo S, Pérez-Pérez G, Alarcón T, López-Brea M (2011) Rapid detection of clarithromycin resistant Helicobacter pylori strains in Spanish patients by polymerase chain reaction-restriction fragment length polymorphism. Rev Esp Quimioter 24: 32-36.
  • Alvarez A, Moncayo JI, Santacruz JJ, Santacoloma M, Corredor LF, Reinosa E (2009) Antimicrobial susceptibility and mutations involved in clarithromycin resistance in Helicobacter pylori isolates from patients in the western central region of Colombia. Antimicrob Agents Chemother 53: 4022-4024.
  • Ando T, Goto Y, Maeda O, Watanabe O, Ishiguro K, Goto H (2006) Causal role of Helicobacter pylori infection in gastric cancer. World J Gastroenterol 12: 181-186.
  • Booka M, Okuda M, Shin K, Miyashiro E, Hayashi H, Yamauchi K, Tamura Y, Yoshikawa N (2005) Polymerase chain reaction - restriction fragment length polymorphism analysis of clarithromycin-resistant Helicobacter pylori infection in children using stool sample. Helicobacter 10: 205-213.
  • Chisholm SA, Owen RJ, Teare EL, Saverymuttu S (2001) PCR-based diagnosis of Helicobacter pylori infection and real-time determination of clarithromycin resistance directly from human gastric biopsy samples. J Clin Microbiol 39: 1217-1220.
  • Dzierżanowska D (2009) Makrolidy i ketolidy. In: Antybiotykoterapia praktyczna, pp 133-140. α-medica press, Bielsko-Biała (in Polish).
  • Erah PO, Goddard AF, Barrett DA, Shaw PN, Spiller RC (1997) The stability of amoxycillin, clarithromycin and metronidazole in gastric juice: relevance to the treatment of Helicobacter pylori infection. J Antimicrob Chemother 39: 5-12.
  • European Committee on Antimicrobial Susceptibility Testing (2013) Breakpoint tables for interpretation of MICs and zone diameters. Version 3.1, valid from 2013-02-11:
  • Figueiredo C, Garcia-Gonzalez MA, Machado JC (2013) Molecular pathogenesis of gastric cancer. Helicobacter 18: 28-33.
  • Francavilla R, Lionetti E, Castellaneta S, Margiotta M, Piscitelli D, Lorenzo L, Cavallo L, Ierardi E (2010) Clarithromycin-resistant genotypes and eradication of Helicobacter pylori. J Pediatr 157: 228-232.
  • de Francesco V, Margiotta M, Zullo A, Hassan C, Valle ND, Burattini O, Cea U, Stoppino G, Amoruso A, Stella F, Morini S, Panella C, Ierardi E (2006) Primary clarithromycin resistance in Italy assessed on Helicobacter pylori DNA sequences by TaqMan real-time polymerase chain reaction. Aliment Pharmacol Ther 23: 429-435.
  • Fujiki S, Iwao Y, Kobayashi M, Miyagishima A, Itai S (2011) Stabilization mechanism of clarithromycin tablets under gastric pH conditions Chem Pharm Bull (Tokyo) 59: 553-558.
  • Gerrits MM, van Vliet AH, Kuipers EJ, Kusters JG (2006) Helicobacter pylori and antimicrobial resistance: molecular mechanisms and clinical implications. Lancet Infect Dis 6: 699-709.
  • Giorgio F, Principi M, De Francesco V, Zullo A, Losurdo G, Di Leo A, Ierardi E (2013) Primary clarithromycin resistance to Helicobacter pylori: Is this the main reason for triple therapy failure? World J Gastrointest Pathophysiol 4: 43-46.
  • Gisbert JP, Calvet X (2011) Review article: common misconceptions in the management of Helicobacter pylori-associated gastric MALT-lymphoma. Aliment Pharmacol Ther 34: 1047-1062.
  • Hao Q, Li Y, Zhang ZJ, Liu Y, Gao H (2004) New mutation points in 23S rRNA gene associated with Helicobacter pylori resistance to clarithromycin in northeast China. World J Gastroenterol 10: 1075-1077.
  • Hirata K, Suzuki H, Nishizawa T, Tsugawa H, Muraoka H, Saito Y, Matsuzaki J, Hibi T (2010) Contribution of efflux pumps to clarithromycin resistance in Helicobacter pylori. J Gastroenterol Hepatol 25 Suppl 1: S75-S79.
  • Ho SL, Tan EL, Sam CK, Goh KL (2010) Clarithromycin resistance and point mutations in the 23S rRNA gene in Helicobacter pylori isolates from Malaysia. J Dig Dis 11: 101-105.
  • den Hollander WJ, Sostres C, Kuipers EJ, Lanas A (2013) Helicobacter pylori and nonmalignant diseases. Helicobacter 18: 24-7.
  • International Agency for Research on Cancer (IARC) (1994) Monograph on the Evaluation of Carcinogenic Risks to Humans, Vol. 61, Schistosomes, Liver Flukes and Helicobacter pylori. IARC, Lyon, France.
  • Karczewska E, Klesiewicz K, Skiba I, Wojtas-Bonior I, Sito E, Czajecki K, Zwolińska-Wcisło M, Budak A (2012) Variability in prevalence of Helicobacter pylori strains resistant to clarithromycin and levofloxacin in Southern Poland. Gastroenterol Res Pract 2012: 418010.
  • Karczewska E, Wojtas I, Budak A (2009) Prevalence of Helicobacter pylori primary resistance to antimicrobial agents in Poland and around the world. Post Mikrobiol 48: 31-41.
  • Karczewska E, Wojtas-Bonior I, Sito E, Zwolińska-Wcisło M, Budak A (2011) Primary and secondary clarithromycin, metronidazole, amoxicillin and levofloxacin resistance to Helicobacter pylori in southern Poland. Pharmacol Rep 63: 799-807.
  • Konturek PC, Konturek SJ, Brzozowski T (2009) Helicobacter pylori infection in gastric cancerogenesis. J Physiol Pharmacol 60: 3-21.
  • Malfertheiner P, Megraud F, O'Morain CA, Atherton J, Axon AT, Bazzoli F, Gensini GF, Gisbert JP, Graham DY, Rokkas T, El-Omar EM, Kuipers EJ (2012) European Helicobacter Study Group. Management of Helicobacter pylori infection - the Maastricht IV/ Florence Consensus Report. Gut 61: 646-664.
  • Mégraud F (2004) Helicobacter pylori antibiotic resistance: prevalence, importance, and advances in testing. Gut 53: 1374-1384.
  • Mégraud F, Coenen S, Versporten A, Kist M, Lopez-Brea M, Hirschl AM, Andersen LP, Goossens H, Glupczynski Y (2013) Study Group participants. Helicobacter pylori resistance to antibiotics in Europe and its relationship to antibiotic consumption. Gut 62: 34-42.
  • Mégraud F, Lehours P (2007) Helicobacter pylori detection and antimicrobial susceptibility testing. Clin Microbiol Rev 20: 280-322.
  • Ménard A, Santos A, Mégraud F, Oleastro M (2002) PCR-restriction fragment length polymorphism can also detect point mutation A2142C in the 23S rRNA gene, associated with Helicobacter pylori resistance to clarithromycin. Antimicrob Agents Chemother 46: 1156-1157.
  • Occhialini A, Urdaci M, Doucet-Populaire F, Bébéar CM, Lamouliatte H, Mégraud 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.
  • Oleastro M, Ménard A, Santos A, Lamouliatte H, Monteiro L, Barthélémy P, Mégraud F (2003) Real-time PCR assay for rapid and accurate detection of point mutations conferring resistance to clarithromycin in Helicobacter pylori. J Clin Microbiol 41: 397-402.
  • Perez-Perez GI, Rothenbacher D, Brenner H (2004) Epidemiology of Helicobacter pylori infection. Helicobacter 9 Suppl 1: 1-6.
  • Raymond J, Burucoa C, Pietrini O, Bergeret M, Decoster A, Wann A, Dupont C, Kalach N (2007) Clarithromycin resistance in Helicobacter pylori strains isolated from French children: prevalence of the different mutations and coexistence of clones harboring two different mutations in the same biopsy. Helicobacter 12: 157-163.
  • Rimbara E, Noguchi N, Yamaguchi T, Narui K, Kawai T, Sasatsu M (2005) Development of a highly sensitive method for detection of clarithromycin-resistant Helicobacter pylori from human feces. Curr Microbiol 51: 1-5.
  • Rożynek E, Dzierżanowska-Fangrat K, Celińska-Cedro D, Jóźwiak P, Madaliński K, Dzierżanowska D (2002) Primary resistance of Helicobacter pylori to antimicrobial agents in Polish Children. Acta Microbiol Pol 51: 255-263.
  • Schabereiter-Gurtner C, Hirschl AM, Dragosics B, Hufnagl P, Puz S, Kovách Z, Rotter M, Makristathis A (2004) Novel real-time PCR assay for detection of Helicobacter pylori infection and simultaneous clarithromycin susceptibility testing of stool and biopsy specimens. J Clin Microbiol 42: 4512-4518.
  • Stone GG, Shortridge D, Versalovic J, Beyer J, Flamm RK, Graham DY, Ghoneim AT, Tanaka SK (1997) A PCR-oligonucleotide ligation assay to determine the prevalence of 23S rRNA gene mutations in clarithromycin-resistant Helicobacter pylori. Antimicrob Agents Chemother 41: 712-714.
  • Taylor DE, 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.
  • Versalovic J, Osato MS, Spakovsky K, Dore MP, Reddy R, Stone GG, Shortridge D, Flamm RK, Tanaka SK, Graham DY (1997) Point mutations in the 23S rRNA gene of Helicobacter pylori associated with different levels of clarithromycin resistance. J Antimicrob Chemother 40: 283-286..
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