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Antibiotic resistance of Helicobacter pylori in Mashhad, Iran
Abolfazl Zendedel,1 Farideh Moradimoghadam,2 Vahid Almasi,3 Hamidreza Zivarifar4 Abstract
Objective: To evaluate Helicobacter pylori resistance to amoxicillin, clarithromycin, metronidazole and tetracycline
in Mashhad, Iran.
Methods: The cross-sectional study was done from January to May 2008 in Mashhad, involving 185 patients who
had been indicated for endoscopy and lesions had been found. Biopsy samples were assessed with histological
evaluation, rapid urease test, and culture. Antibiotic resistance was assessed by the disc diffusion method. Data was
analysed with SPSS 11.5 using chi-square and Fisher’s exact test. P values of < 0.05 were regarded as statistically
significant.
Results: Of the total patients, histological evaluations were positive in 124 (67%). Compared with histology,
sensitivity and specificity of rapid urease test were 96.7% and 100%, respectively. In 82 (66.1%) patients with positive
cultures, antibiotic resistance was found in 14 (17.1%) for clarithromycin; 53 (64.6%) for metronidazole; and 8 (9.8%)
for amoxicillin. No resistance was observed for tetracycline. Moreover, 9 (64%) patients with resistance to
clarithromycin had co-resistance to metronidazole.
Conclusion: Metronidazole is not recommended for treatment of Helicobacter pylori as a first-line drug. Also,
considering the sensitivity and specificity of rapid urease test, we suggest this method as a suitable alternative for
histology.
Keywords: Helicobacter pylori, Antibiotic resistance, Antibiotic. (JPMA 63: 336; 2013)
Introduction
compared with that in 2005.15 Also, Sun et al reported that Helicobacter pylori (H. pylori) has been reported as the in Shanghai the resistance to clarithromycin increased cause of some frequent gastrointestinal disorders.1,2 from 8.6% in 2000 to 20.7% in 2009.16 Furthermore, Gao et Gastroduodenal ulcer disease and gastric malignancies al. declared that H. pylori resistance to metronidazole, are caused by H. pylori.2-6 This bacterium has been clarithromycin and fluoroquinolone increased from 2000 classified as a type-1 carcinogen in the World Health Organization report.7 Infection with H. pylori is so The aim of this study was to evaluate H. pylori resistance common that more than 50% of the world's population isinfected with this bacterium.8 This bacterium is usually to amoxicillin, clarithromycin, metronidazole and acquired in childhood.2 Eradication of H. pylori has tetracycline in Mashhad city of Iran.
prophylactic effect on gastric malignancies.5 Patients and methods
Several combined antibacterial regimens have been The cross-sectional study was conducted from January to advised for the eradication of H. pylori. In European May 2008 at the Gastrointestinal Clinic of Ghaem Hospital, guidelines, clarithromycin and either amoxicillin or Mashhad, northeastern Iran. Patients with chronic metronidazole are the first-line antibiotics.3 Also, in other dyspepsia, who had been indicated for endoscopy and advised regimens tetracycline has been suggested.5,9 had lesions in their endoscopy, were enrolled in the study.
Current anti-H. pylori therapies can fail in up to 30% of We did not intend to do endoscopy in patients with no patients.3,5 Antibiotic resistance is an essential cause of indication for endoscopy, thus we selected these patients.
Patients with a history of using 3 or 4 drugs for thetreatment of H. Pylori, those with gastrointestinal Over time, antibiotic resistance of H. pylori has malignancy, or those with recent (2 weeks before the increased.13,14 Siavoshi et al reported that resistance to evaluation) use of antibiotics, bismuth or proton pomp metronidazole and tetracycline increased in 2008 inhibitors (PPIs) were excluded because antibiotics andsome other drugs can impair the results of sensitivity 1,3Lorestan University of Medical Sciences, 2,4Mashhad University of Medical tests. Indications of endoscopy in the patients with chronic dyspepsia included incomplete treatment with Correspondence: Farideh Moradimoghadam. EMail: [email protected]
antacid drugs, presence of suspicious signs (weight loss, Antibiotic resistance of Helicobacter pylori in Mashhad, Iran anaemia of undetermined cause, gastrointestinal After the surface of the plates got dried, the antibiotic discs were transferred to the surface of the plates. After 15 consistent vomiting and palpable abdominal mass), age over 45 years, history of heartburn for more than 5 years, microaerophilic condition and they were kept in this and being suspicious of cancer or organic diseases. The condition at 37° C. After 5 days, the inhibition zone patients were divided into 3 age groups: <30 years; 30-50 diametre (IZD) was recorded. IZDs more than or equal to years; and >50 years. The study was approved by the 14, 15, 17 and 20mm were considered to be susceptible to Research and Ethical Committee of Mashhad University of Uring consecutive sampling, the sample size was Data was analysed with SPSS 11.5, using chi-square and Fisher’s exact test. P values of <0.05 were regarded asstatistically significant.
Results
A total of 185 individuals were enrolled in the study. Of
them, 101 (54.6%) were female; histological evaluations
were positive in 124 (67%); and 65 (52.4%) of them were
All the endoscopies were done by an Internal Medicine female. There was no relationship between gender and H.
physician. The samples were taken from the antrum of the stomach. A part of each sample was specified for rapidurease test (RUT) and another part was specified for The mean age of the patients was 41.3±15.2 years; For RUT, the samples were transferred to a sterile urease Infection with H. pylori was more common in the older test broth environment. Phenol red indicator was then patients. There was a significant relationship between age added. To assess the change of the samples colour, they and infection with H. pylori (p <0.012) (Table-1).
were kept at 37°C and were observed for 1 hour. When the Compared with histology, sensitivity and specificity of sample colour changed from yellow to pink, the result of RUT were 96.7% and 100%, and sensitivity and specificity of culture were 66.1% and 100%, respectively.
The biopsy samples were transferred to the laboratory for In 82 patients with positive cultures, antibiotic resistance culture and staining in semi-solid (0.1% agar) normal was found in 14 (17.1%) patients for clarithromycin; 53 saline. The samples were prepared for Gram staining and (64.6%) patients for metronidazole; and 8 (9.8%) for amoxicillin. No resistance was observed for tetracycline. For culturing the samples, 7% foetal calf serum and 10% There was no relationship between age and resistance to lysed horse blood and the antibiotics (vancomycin clarithromycin, amoxicillin or tetracycline (p<0.05).
(10µg/L), amphotericin B (5µg/L) and trimethoprim However, resistance to metronidazole was more common in (5µg/L)) were added to RUT media and Colombia agar patients younger than 50 years of age (p<0.010) (Table-2). base (Merck, Germany). The samples were then placed onit. It was incubated at 37°C for 3-5 days under There was no significant relationship between antibiotic microaerophilic conditions. H. Pylori was identified as gram-negative with spiral or curved rods, which produce In addition, resistance to 3 antibiotics (clarithromycin, urease, catalase and oxidase. Sensitivity and specificity ofRUT and culture were compared with histology as the Table-1: Distribution of patients and positive histology in age groups.
Percentage of
In cases with positive cultures, antibiotic resistance was patients
patients with
patientswith
assessed by using the disc diffusion method for positive histology
positive histology
tetracycline. Brain heart infusion broth (Merck, Germany) plates were used with added 10% foetal calf serum.
Bacterial suspension (equivalent to Mac Farland no. 3 turbidity) was transferred on the surface of the plates.
A. Zendedel, F. Moradimoghadam, V. Almasi, et al.
Table-2: The frequency of resistance to clarithromycin, metronidazole and amoxicillin in age groups.
<30 years
30-50 years
>50 years
P-value*
Number of patients with positive cultures * Calculated using the chi-square test.
metronidazole and amoxicillin) was seen in 1 (1.2%) 17.1% for clarithromycin. In other studies, resistance to patient, 21 (25.6%) patients had no resistance to any of clarithromycin was between 7.3% in Iran from 2005 to the 4 antibiotics; and 9 (64%) patients with resistance to 200815 and 44% in Iran between 2008 and 2009.21 In Iran, clarithromycin had co-resistance to metronidazole.
it was reported to be 30% in the north from 2007 to2010,20 44% in the south between 2008 and 200921 and Discussion
7.3% in the center from 2005 to 2008.15 In developing H. pylori resistance to antibiotics is a worldwide problem.
countries, it was reported to be 14.6% in Tunisia from 2005 Over time, H. pylori resistance to antibiotics is to 2007.13 In developed countries, it was declared to be increasing.13,14 Considering the role of this bacterium in 20.7% in Shanghai (China) in 2009,16 13.2% in Ireland from some frequent gastrointestinal disorders (such as 2007 to 200819 and 26% in France between 2004 and gastroduodenal ulcer disease) and gastric malignancies, 2007.6 Resistance to clarithromycin in our study was lower determination of antibiotic resistance to H. pylori is an than other studies that were done in the north and the south of Iran, but higher than the study carried out in the A total of 185 individuals were enrolled in this study, and centre of Iran. Also, it is similar to the studies that were 67% of them were positive in terms of histological done in Tunisia, Shanghai (China) and Ireland.
evaluations. In our study, there was no relationship In our study, the antibiotic resistance was found to be between gender and infection. However, in Naja et al.'s 9.8% for amoxicillin. In other studies, resistance to study, infection with H. pylori was more common in amoxicillin ranged from no resistance in Shanghai in 2009,16 in France from 2004 to 20076 and in Tunisia from In the present study, infection with H. pylori was more 2005 to 2007,13 to 20% in Iran between 2008 and 2009.21 common in the older patients. This finding is consistent In Iran, it was reported to be 6.8% in the north from 2007 to 2010,20 20% in the south between 2008 and 200921 and7.3% in the centre from 2005 to 2008.15 Resistance to Compared with histology, sensitivity and specificity of amoxicillin in our study was similar to other studies that RUT were 96.7% and 100%, and sensitivity and specificity were done in the north and the centre of Iran, but lower of culture were 66.1% and 100%, respectively. According than the study done in the south of Iran.
to these findings, the use of RUT as an alternative methodseems to be logical. In our study, no resistance was observed for tetracycline.
In other studies, resistance to tetracycline ranged from no In our study, the antibiotic resistance was found to be resistance in France between 2004 and 20076 to 38.1% in 64.6% for metronidazole, which was in line with earlier Iran from 2005 to 2008.15 In Iran, it was reported to be 9% findings. Resistance to metronidazole ranged from 31.5% in the north from 2007 to 2010,20 3% in the south in Ireland from 2007 to 200819 to 95.5% in South Africa in between 2008 and 200921 and 38.1% in the centre from 2009.11 In Iran, it was reported to be 73.4% in the north 2005 to 2008.15 Resistance to tetracycline in this study is from 2007 to 2010,20 44% in the south between 2008 and lower than the studies performed in Iran. It is massively 2009,21 and 55.6% in the centre from 2005 to 2008.15 In different from the study that was done in the centre of developing countries, it was reported to be 95.5% in Iran. It is similar to the amount of resistance to tetracycline South Africa in 2009,11 and 56.8% in Tunisia from 2005 to 2007.13 In developed countries, it was declared to be 50% In this study, there was no relationship between age and in Shanghai (China) in 2009,16 31.5% in Ireland from 2007 resistances to clarithromycin, amoxicillin or tetracycline, to 2008,19 and 61% in France between 2004 and 2007.6 but resistance to metronidazole was more common in the In our study, the antibiotic resistance was found to be patients younger than 50 years age (p<0.05). An earlier Antibiotic resistance of Helicobacter pylori in Mashhad, Iran study reported no relationship between age and Vietnam: a cross-sectional, hospital-based study. BMC Furuta T, Graham DY. Pharmacologic aspects of eradication In the present study, there was no significant difference therapy for Helicobacter pylori Infection. Gastroenterol Clin NorthAm 2010; 39: 465-80.
between antibiotic resistance of H. pylori and gender. It is Romano M, Cuomo A, Gravina AG, Miranda A, Iovene MR, Tiso A, et consistent with literature,12 but there has been one study al. Empirical levofloxacin-containing versus clarithromycin- reporting resistance to metronidazole being higher in containing sequential therapy for Helicobacter pylori eradication: a randomised trial. Gut 2010; 59: 1465-70.
Tanih NF, Okeleye BI, Naidoo N, Clarke AM, Mkwetshana N, Green Conclusion
E, et al. Marked susceptibility of South African Helicobacter pyloristrains to ciprofloxacin and amoxicillin: clinical implications. S Afr Considering the high resistance to metronidazole, it is not recommended as a first-line drug. Using clarithromycin, Boyanova L, Ilieva J, Gergova G, Spassova Z, Nikolov R, Davidkov L, amoxicillin and tetracycline in the H. pylori treatment et al. Evaluation of clinical and socio-demographic risk factors for regimen seems to be logical. Moreover, according to the antibacterial resistance of Helicobacter pylori in Bulgaria. J MedMicrobiol 2009; 58: 94-100.
sensitivity and specificity of RUT, it can be a suitable Ben Mansour K, Burucoa C, Zribi M, Masmoudi A, Karoui S, Kallel L, et al. Primary resistance to clarithromycin, metronidazole andamoxicillin of Helicobacter pylori isolated from Tunisian patients Acknowledgments
with peptic ulcers and gastritis: a prospective multicentre study.
The authors are grateful to Mr. Yadollah Pournia, instructor Ann Clin Microbiol Antimicrob 2010; 9: 22.
De Francesco V, Giorgio F, Hassan C, Manes G, Vannella L, Panella of English language at Lorestan University of Medical C, et al. Worldwide H. pylori antibiotic resistance: a systematic Sciences, and the Clinical Research Centre of Lorestan review. J Gastrointestin Liver Dis 2010; 19: 409-14.
Siavoshi F, Saniee P, Latifi-Navid S, Massarrat S, Sheykholeslami A.
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