The mutation of the rdxA gene in metronidazole-resistant Helicobacter pylori clinical isolates

Authors

1 Department of Biology, Tonekabon Branch, Islamic Azad University, Tonekabon, Iran

2 Department of Microbiology, Isfahan University of Medical Sciences, Isfahan, Iran

3 Department of Food Hygiene, Azad University of Shahrekord, Shahrekord, Iran

Abstract

Backgrounds: Antibiotic resistance is an increasing problem throughout the developed world, and knowledge about different resistance mechanisms is consequential for efficient treatment of bacterial infections. Although metronidazole has been frequently used in treatment regimens for H. pylori infection, but antibiotic resistance is now a major contributing factor in treatment failure. Nevertheless metronidazole has been greatly used as a critical component of combination therapies for H. pylori infection.
Objective: This study is trying to describe the mutational mechanisms of metronidazole resistance in H. pylori in our clinical isolates in Isfahanian patients, Iran and compare with the findings of previous studies in world.
Materials and Methods: MIC values of metronidazole for H. pylori strains were determined by E- test. Both rdxA and glmM genes used for confirmation of isolates as H. pylori and then amplification of another rdxA oligonucleotide pair was done. Finally, the six resistant strains were sent to sequencing for other processing and further analysis was done by software.
Results: The result of six clinical isolates in comparison with 26695, J99 and 69A as a sensitive and resistant reference strains showed plenty of mutations. No frame shift and nonsense mutation was seen in our clinical isolates.
Conclusion: An interesting finding in metronidazole-resistant strains in our study was the detection of one mutation not previously described in the literature in the rdxA gene and this W(209)R substitution presumably plays a role in inducing metronidazole resistance.

Keywords

1. Leiros HK, Kozielski-Stuhrmann S, Kapp U, Terradot L, Leonard GA, McSweeney SM. Structural Basis of 5-Nitroimidazole Antibiotic Resistance. J Biol Chem 2004;279:55840-9.  Back to cited text no. 1
    
2. Han F, Liu S, Ho B, Yan Z, Yan X. Alterations in rdxA and frxA genes and their upstream regions in metronidazole-resistant Helicobacter pylori isolates. Res Microbiol 2007;158:38-44.   Back to cited text no. 2
    
3. Buta N, Tanih NF, Ndip RN. Increasing trend of metronidazole resistance in the treatment of Helicobacter pylori infection: A global challenge. Afr J Biotechnol 2010;9:1115-21.   Back to cited text no. 3
    
4. Diniz CG, Farias LM, Carvalho MA, Rocha ER, Smith CJ. Differential gene expression in a Bacteroides fragilis metronidazole-resistant mutant. J Antimicrob Chemother 2004;54:100-8.   Back to cited text no. 4
    
5. Jenks PJ, Edwards DL. Metronidazole resistance in Helicobacter pylori. Int J Antimicrob Agents 2002;19:1-7.  Back to cited text no. 5
    
6. Lofmark S, Edlund C, Nord CE. Metronidazole is still the drug of choice for treatment of anaerobic infections. Clin Infect Dis 2010;50:S16-23.  Back to cited text no. 6
    
7. Yang YJ, Wui J, Sheu BS, Kao AW, Huang AH. The rdxA gene plays a more major role than frxA gene mutation in high level metronidazole resistance of helicobacter pylori in Taiwan. Helicobacter J 2004;9:400-7.   Back to cited text no. 7
    
8. Matteo MJ, Perez CV, Domingo MR, Olmos M, Sanchez C, Catalano M. DNA sequence analysis of rdxA and frxA from paired metronidazole-sensitive and -resistant Helicobacter pylori isolates obtained from patients with heteroresistance. Int J Antimicrob Agents 2006;27:152-8.   Back to cited text no. 8
    
9. Chisholm SA, Owen RJ. Mutations in Helicobacter pylori rdxA gene sequences may not contribute to metronidazole resistance. J Antimicrob Chemother 2003;51:995-9.   Back to cited text no. 9
    
10. Mirzaei N, Poursina F, Faghri J, Talebi M, Khataminezhad MR, Hasanzadeh A, Ghasemian Safaei H. Prevalence of resistance to Helicobacter pylori strains to selected antibiotics in Isfahan, Iran. Jundishapur J Microbiol 2013;6:e6342.  Back to cited text no. 10
    
11. Cordova Espinoza MG, Vazquez RG, Mendez IM, Vargas CR, Cerezo SG. Detection of the glmM Gene in Helicobacter pylori Isolates with a Novel Primer by PCR. J Clin Microbiol 2011;49:1650-2.   Back to cited text no. 11
    
12. Jeong JY, Mukhopadhyay A, Akada JK, Dailidiene D, Hoffman PS, Berg DE. Roles of FrxA and RdxA Nitroreductases of Helicobacter pylori in Susceptibility and Resistance to Metronidazole. J Bacteriol 2001;183:5155-62.   Back to cited text no. 12
    
13. Ossenkopp YJ, Pot R, Westerloo DJ, Goodwin A, Grauls CV, Berg DE, et al. Insertion of Mini-IS605 and Deletion of Adjacent Sequences in the Nitroreductase (rdxA) Gene Cause Metronidazole Resistance in Helicobacter pylori NCTC11637. Antimicrob Agents Chemother 1999;43:2657-62.   Back to cited text no. 13
    
14. Kargar M, Baghernejad M, Doosti A. Role of NADPH-insensitive nitroreductase gene to metronidazole resistance of Helicobacter pylori strains. Daru 2010;18:137-40.  Back to cited text no. 14
    
15. Abdollahi H, Savari M, Zahedi MJ, Moghadam SD, Abasi MH. A study of rdxA gene deletion in metronidazole resistant and sensitive Helicobacter pylori isolates in Kerman, Iran. Jundishapur J Microbiol 2010;4:99-104.  Back to cited text no. 15
    
16. Mohammadi M, Doroud D, Mohajerani N, Massarrat S. Helicobacter pylori antibiotic resistance in Iran. World J Gastroenterol 2005;11:6009-13.  Back to cited text no. 16
    
17. Jeong JY, Mukhopadhyay AK, Dailidiene D, Wang Y, Velapatino B, Gilman RH, et al. Sequential Inactivation of rdxA (HP0954) and frxA (HP0642) Nitroreductase Genes Causes Moderate and High-Level Metronidazole Resistance in Helicobacter pylori. J Bacteriol 2000;182:5082-90.  Back to cited text no. 17
    
18. Llanes R, Soria C, Nagashima S, Kobayashi N, Gala A, Guzman D, et al. Phenotypic and Genetic Characterization of Antimicrobial Profiles of Helicobacter pylori Strains in Cuba. J Health Popul Nutr 2010;28:124-9.  Back to cited text no. 18
    
19. Kim SY, Joo YM, Lee HS, Chung IS, Yoo YJ, Merrell DS, et al. Genetic analysis of Helicobacter pylori clinical isolates suggests resistance to metronidazole can occur without the loss of functional rdxA. J Antibiot 2009;62:43-50.   Back to cited text no. 19
    
20. Known DH, Pena JA, Osato MS, Fox JG, Graham DY, Versalovic J. Frameshift mutations in rdxA and metronidazole resistance in North American Helicobacter pylori isolates. J Antimicrob Chemother 2000;46:793-6.  Back to cited text no. 20
    
21. Solca NM, Bernasconi MR, Piffaretti JC. Mechanism of Metronidazole Resistance in Helicobacter pylori: Comparison of the rdxA Gene Sequences in 30 Strains. Antimicrob Agents Chemother 2000;44:2207-10.  Back to cited text no. 21
    
22. Latham SR, Owen RJ, Elviss NC, Labigne A, Jenks PJ. Differentiation of metronidazole-sensitive and -resistant clinical isolates of helicobacter pylori by immunoblotting with antisera to the RdxA protein. J Clin Microbiol 2001;39:3052-5.  Back to cited text no. 22
    
23. Bereswill S, Krainick C, Stahler F, Herrmann L, Kist M. Analysis of the rdxA gene in high-level metronidazole-resistant clinical isolates confirms a limited use of rdxA mutations as a marker for prediction of metronidazole resistance in Helicobacter pylori. FEMS Immunol Med Microbiol 2003;36:193-8.  Back to cited text no. 23