Antimicrobial susceptibility and clarithromycin resistance patterns of Helicobacter pylori clinical isolates in Vietnam


Helicobacter pylori is a gastric pathogen that causes several gastroduodenal disorders such as peptic ulcer disease and gastric cancer. Eradication efforts of H. pylori are often hampered by antimicrobial resistance in many countries, including Vietnam. Here, the study aimed to investigate the occurrence of antimicrobial resistance among H. pylori clinical isolates across 13 hospitals in Vietnam. The study further evaluated the clarithromycin resistance patterns of H. pylori strains. In order to address the study interests, antimicrobial susceptibility testing, epsilometer test and PCR-based sequencing were performed on a total of 193 strains isolated from patients, including 136 children (3–15 years of age) and 57 adults (19–69 years of age). Antimicrobial susceptibility testing showed that the overall resistance to amoxicillin, clarithromycin, levofloxacin, metronidazole, and tetracycline was 10.4%, 85.5%, 24.4%, 37.8%, and 23.8% respectively. The distribution of minimum inhibitory concentrations (MICs) of clarithromycin-resistant strains was 85.5% with MIC >0.5 μg/mL. The majority of the clarithromycin resistant isolates (135 of 165 subjects) have MICs ranging from 2 μg/mL to 16 μg/mL. Furthermore, sequencing detection of mutations in 23S rRNA gene revealed that strains resistant and susceptible to clarithromycin contained both A2143G and T2182C mutations. Of all isolates, eight clarithromycin-resistant isolates (MIC >0.5 μg/mL) had no mutations in the 23S rRNA gene. Collectively, these results demonstrated that a proportion of clarithromycin-resistant H. pylori strains, which are not related to the 23S rRNA gene mutations, could be potentially related to other mechanisms such as the presence of an efflux pump or polymorphisms in the CYP2C19 gene. Therefore, the present study suggests that providing susceptibility testing prior to treatment or alternative screening strategies for antimicrobial resistance is important for future clinical practice. Further studies on clinical guidelines and treatment efficacy are pivotal for successful eradication of H. pylori infection.


Suerbaum S, Michetti P. (2002). Helicobacter pylori infection. N Engl J Med. 347(15): 1175–1186.

Peek RM Jr, Blaser MJ. (2002). Helicobacter pylori and gastrointestinal tractadenocarcinomas. Nat Rev Cancer. 2(1): 28–37.

Lacy BE, Rosemore J. (2001). Helicobacter pylori: ulcers and more: the beginning of anera. J Nutr. 131(10): 2789S–2793S.

Eusebi LH, Zagari RM, Bazzoli F. (2014). Epidemiology of Helicobacter pylori infection.Helicobacter. 19(Suppl 1): 1–5.

Hoang TT, Bengtsson C, Phung DC, et al. (2005). Seroprevalence of Helicobacter pyloriinfection in urban and rural Vietnam. ClinDiagn Lab Immunol. 12(1): 81–85.

Nguyen BV, Nguyen KG, Phung CD, et al. (2006). Prevalence of and factors associatedwith Helicobacter pylori infection in children in the north of Vietnam. Am J TropMed Hyg. 74(4): 536–539.

Nguyen VB, Nguyen GK, Phung DC, et al. (2006). Intra-familial transmission of Helicobacterpylori infection in children of households with multiple generations in Vietnam. Eur J Epidemiol. 21(6): 459–463.

Hosking SW, Ling TK, Chung SC, et al. (1994). Duodenal ulcer healing by eradicationof Helicobacter pylori without anti-acid treatment: randomised controlled trial. Lancet. 343(8896): 508–510.

Takenaka R, Okada H, Kato J, et al. (2007). Helicobacter pylori eradication reduced theincidence of gastric cancer, especially of the intestinal type. Aliment PharmacolTher. 25(7): 805–812.

Malfertheiner P: Helicobacter pylori infection--management from a Europeanperspective. Dig Dis. 2014; 32(3): 275–280.

Malfertheiner P, Megraud F, O'Morain CA, et al.: Management of Helicobacterpylori infection--the Maastricht IV/ Florence Consensus Report. Gut. 2012; 61(5): 646–664.

Selgrad M, Bornschein J, Malfertheiner P: Guidelines for treatment of Helicobacterpylori in the East and West. Expert Rev Anti Infect Ther. 2011; 9(8): 581–588.

Fock KM, Katelaris P, Sugano K, et al.: Second Asia-Pacific ConsensusGuidelines for Helicobacter pylori infection. J GastroenterolHepatol. 2009; 24(10): 1587–1600.

Chey WD, Wong BC: Practice Parameters Committee of the American College of Gastroenterology: American College of Gastroenterology guideline on themanagement of Helicobacter pylori infection. Am J Gastroenterol. 2007; 102(8): 1808–1825.

Jenks PJ: Causes of failure of eradication of Helicobacter pylori. BMJ. 2002; 325(7354): 3–4.

Vakil N, Megraud F: Eradication therapy for Helicobacter pylori. Gastroenterology. 2007; 133(3): 985–1001.

Mégraud F: H pylori antibiotic resistance: prevalence, importance, andadvances in testing. Gut. 2004; 53(9): 1374–1384.

Binh TT, Shiota S, Nguyen LT, et al.: The incidence of primary antibiotic resistanceof Helicobacter pylori in Vietnam. J ClinGastroenterol. 2013; 47(3): 233–238.

Phan TN, Santona A, Tran VH, et al.: High rate of levofloxacin resistance in abackground of clarithromycin- and metronidazole-resistant Helicobacter pyloriin Vietnam. Int J Antimicrob Agents. 2015; 45(3): 244–248.

Nguyen TV, Bengtsson C, Yin L, et al.: Eradication of Helicobacter pylori inchildren in Vietnam in relation to antibiotic resistance. Helicobacter. 2012; 17(4): 319–325.

Versalovic J, Shortridge D, Kibler K, et al.: Mutations in 23S rRNA are associatedwith clarithromycin resistance in Helicobacter pylori. Antimicrob AgentsChemother. 1996; 40(2): 477–480.

Lee HJ, Kim JI, Cheung DY, et al.: Eradication of Helicobacter pylori accordingto 23S ribosomal RNA point mutations associated with clarithromycinresistance. J Infect Dis. 2013; 208(7): 1123–1130.

Francesco VD, Zullo A, Hassan C, et al.: Mechanisms of Helicobacter pyloriantibiotic resistance: An updated appraisal. World J GastrointestPathophysiol2011; 2(3): 35–41.

Mégraud F, Lehours P: Helicobacter pylori detection and antimicrobialsusceptibility testing. ClinMicrobiol Rev. 2007; 20(2): 280–322.

Kim KS, Kang JO, Eun CS, et al.: Mutations in the 23S rRNA gene of Helicobacterpylori associated with clarithromycin resistance. J Korean Med Sci. 2002; 17(5): 599–603.

Taylor DE, Ge Z, Purych D, et al.: Cloning and sequence analysis of two copiesof a 23S rRNA gene from Helicobacter pylori and association of clarithromycinresistance with 23S rRNA mutations. Antimicrob Agents Chemother. 1997; 41(12): 2621–2628.

Versalovic J, Osato MS, Spakovsky K, et al.: Point mutations in the 23S rRNAgene of Helicobacter pylori associated with different levels of clarithromycinresistance. J AntimicrobChemother. 1997; 40(2): 283–286.

García-Arata MI, Baquero F, de Rafael L, et al.: Mutations in 23S rRNA inHelicobacter pylori conferring resistance to erythromycin do not always conferresistance to clarithromycin. Antimicrob Agents Chemother. 1999; 43(2): 374–376.

Burucoa C, Landron C, Garnier M, et al.: T2182C mutation is not associated withclarithromycin resistance in Helicobacter pylori. Antimicrob Agents Chemother. 2005; 49(2): 868–870.

Rimbara E, Noguchi N, Kijima H, et al.: Mutations in the 23S rRNA gene ofclarithromycin-resistant Helicobacter pylori from Japan. Int J Antimicrob Agents. 2007; 30(3): 250–254.

Gerrits MM, van Vliet AH, Kuipers EJ, et al.: Helicobacter pylori and antimicrobialresistance: molecular mechanisms and clinical implications. Lancet Infect Dis. 2006; 6(11): 699–709.

Cammarota G, Martino A, Pirozzi G, et al.: High efficacy of 1-week doxycyclineand amoxicillin-based quadruple regimen in a culture-guided, third-linetreatment approach for Helicobacter pylori infection. Aliment PharmacolTher. 2004; 19(7): 789–795.

Perez-Perez GI: Accurate diagnosis of Helicobacter pylori. Culture, includingtransport. GastroenterolClin North Am. 2000; 29(4): 879–884.

European Committee on Antimicrobial Susceptibility Testing: Breakpoint tablesfor interpretation of MICs and zone diameters. Version 4.0. European Society ofClinical Microbiology and Infectious Diseases. 2014.

Tamura K, Peterson D, Peterson N, et al.: MEGA5: molecular evolutionarygenetics analysis using maximum likelihood, evolutionary distance, andmaximum parsimony methods. MolBiolEvol. 2011; 28(10): 2731–2739.

Benson DA, Cavanaugh M, Clark K, et al.: GenBank. Nucleic Acids Res. 2013; 41(Database issue): D36–42.

Graham DY, Fischbach L: Helicobacter pylori treatment in the era of increasingantibiotic resistance. Gut. 2010; 59(8): 1143–1153.

Everhart JE: Recent developments in the epidemiology of Helicobacter pylori.GastroenterolClin North Am. 2000; 29(3): 559–578.

McColl KE: Clinical practice. Helicobacter pylori infection. N Engl J Med. 2010; 362(17): 1597–1604.

Nishizawa T, Maekawa T, Watanabe N, et al.: Clarithromycin Versus Metronidazoleas First-line Helicobacter pylori Eradication: A Multicenter, Prospective,Randomized Controlled Study in Japan. J ClinGastroenterol. 2015; 49(6): 468–471.

Megraud F, Coenen S, Versporten A, et al.: Helicobacter pylori resistance toantibiotics in Europe and its relationship to antibiotic consumption. Gut. 2013; 62(1): 34–42.

Giorgio F, Principi M, De Francesco V, et al.: Primary clarithromycin resistanceto Helicobacter pylori: Is this the main reason for triple therapy failure? World JGastrointestPathophysiol. 2013; 4(3): 43–46.

Ferlay J, Shin HR, Bray F, et al.: Estimates of worldwide burden of cancer in2008: GLOBOCAN 2008. Int J Cancer. 2010; 127(12): 2893–2917.

Nguyen TL, Uchida T, Tsukamoto Y, et al.: Helicobacter pylori infection andgastroduodenal diseases in Vietnam: a cross-sectional, hospital-based study.BMC Gastroenterol. 2010; 10: 114.

Katelaris PH: Helicobacter pylori: antibiotic resistance and treatment options.J GastroenterolHepatol. 2009; 24(7): 1155–1157.

Gonzales R, Bartlett JG, Besser RE, et al.: Principles of appropriate antibioticuse for treatment of acute respiratory tract infections in adults: background,specific aims, and methods. Ann Intern Med. 2001; 134(6): 479–486.

Selgrad M, Tammer I, Langner C, et al.: Different antibiotic susceptibility betweenantrum and corpus of the stomach, a possible reason for treatment failure ofHelicobacter pylori infection. World J Gastroenterol. 2014; 20(43): 16245–16251.

Ayala G, Galvan-Portillo M, Chihu L, et al.: Resistance to antibiotics andcharacterization of Helicobacter pylori strains isolated from antrum and bodyfrom adults in Mexico. Microb Drug Resist. 2011; 17(2): 149–155.

Rimbara E, Noguchi N, Tanabe M, et al.: Susceptibilities to clarithromycin,amoxycillin and metronidazole of Helicobacter pylori isolates from the antrumand corpus in Tokyo, Japan, 1995-2001. ClinMicrobiol Infect. 2005; 11(4):307–311.

Borody TJ, Clancy R, Warren EF, et al.: Antibiotic sensitivities of Helicobacterpylori vary at different gastric mucosal sites. Dordrecht, London, KluwerAcademic. 2003; 373–381.

Gustavson LE, Kaiser JF, Edmonds AL, et al.: Effect of omeprazole onconcentrations of clarithromycin in plasma and gastric tissue at steady state.Antimicrob Agents Chemother. 1995; 39(9): 2078–2083.

Kim JM, Kim JS, Jung HC, et al.: Distribution of antibiotic MICs for Helicobacterpylori strains over a 16-year period in patients from Seoul, South Korea.Antimicrob Agents Chemother. 2004; 48(12): 4843–4847.

De Francesco V, Margiotta M, Zullo A, et al.: Clarithromycin-resistant genotypesand eradication of Helicobacter pylori. Ann Intern Med. 2006; 144(2): 94–100.

van Doorn LJ, Glupczynski Y, Kusters JG, et al.: Accurate prediction of macrolideresistance in Helicobacter pylori by a PCR line probe assay for detectionof mutations in the 23S rRNA gene: multicenter validation study. AntimicrobAgents Chemother. 2001; 45(5): 1500–1504.

Owen RJ: Molecular testing for antibiotic resistance in Helicobacter pylori. Gut.2002; 50(3): 285–289.

Hulten K, Gibreel A, Skold O, et al.: Macrolide resistance in Helicobacter pylori:mechanism and stability in strains from clarithromycin-treated patients.Antimicrob Agents Chemother. 1997; 41(11): 2550–2553.

Occhialini A, Urdaci M, Doucet-Populaire F, et al.: Macrolide resistance inHelicobacter pylori: rapid detection of point mutations and assays of macrolidebinding to ribosomes. Antimicrob Agents Chemother. 1997; 41(12): 2724–2728.

Binh TT, Shiota S, Suzuki R, et al.: Discovery of novel mutations forclarithromycin resistance in Helicobacter pylori by using next-generationsequencing. J AntimicrobChemother. 2014; 69(7): 1796–1803.

Rimbara E, Noguchi N, Kawai T, et al.: Novel mutation in 23S rRNA that conferslow-level resistance to clarithromycin in Helicobacter pylori. Antimicrob AgentsChemother. 2008; 52(9): 3465–3466.

Liu ZQ, Zheng PY, Yang PC: Efflux pump gene hefA of Helicobacter pylori playsan important role in multidrug resistance. World J Gastroenterol. 2008; 14(33):5217–5222.

Hirata K, Suzuki H, Nishizawa T, et al.: Contribution of efflux pumps toclarithromycin resistance in Helicobacter pylori. J GastroenterolHepatol. 2010;25(Suppl 1): S75–79.

Zhang M: High antibiotic resistance rate: A difficult issue for Helicobacterpylori eradication treatment. World J Gastroenterol. 2015; 21(48): 13432–13437.

Quek C, Pham ST, Tran KT, et al.: Dataset 1 in: Antimicrobial susceptibility andclarithromycin resistance patterns of Helicobacter pylori clinical isolates inVietnam. F1000Research. 2016.