In adults as well as in children, several studies have shown that rates of eradication of Helicobacter pylori (H pylori) with the 7-day triple therapy are declining and presently reach only 70% to 80% (1–4). Compared with a triple therapy regimen, higher eradication rates were recently reported with sequential treatment in adults (5–8) and in children (9–13) combining a first association of a proton pump inhibitor (PPI) and amoxicillin (AMO) for the initial 5 days followed by PPI plus clarithromycin (CLA) and metronidazole (MET) for the remaining 5 days. It has been suggested that sequential therapy may overcome CLA resistance, but present data are still not conclusive (14). The rationale for the use of a sequential therapy is that AMO could reduce the bacterial load and destroy the bacterial cell wall. Subsequently, the intracellular diffusion of CLA could be increased with an improved effect with fewer adverse effects than a standard triple therapy (15).
First-line eradication regimens are most often prescribed without tailoring them to antimicrobial susceptibility, although bacterial resistance and low compliance are the main factors for treatment failure (1,13,16). In a European pediatric multicenter study, CLA-resistant and MET-resistant H pylori strains were detected in 24% and 25% of cases, respectively (16). CLA-resistant H pylori strains represent the main cause of treatment failure, particularly in children, MET also being responsible but to a lesser extent (1,15). Prescribing an antibiotic for H pylori eradication based on susceptibility testing is an approach that has been used clinically, allowing “tailored treatment” with marked improvements in treatment success. Indeed, high eradication rates have been obtained by tailoring the triple therapy to the resistance pattern of H pylori(17,18).
As a consequence, the sequential and the “tailored” triple therapy are described as the 2 most effective regimens, but they have not been compared. The purpose of our study was to assess the eradication rate of the sequential treatment compared with a tailored 7-day triple therapy prescribed according to the antimicrobial susceptibility test. The present study allowed us to evaluate the effect of H pylori antimicrobial susceptibility on the eradication rate obtained with the sequential treatment.
PATIENTS AND METHODS
The present prospective, open-label study was conducted between October 2007 and June 2009 in 3 university centers located in Belgium (Brussels), France (Lille), and Italy (Novara). The diagnosis of H pylori infection was based on histology and culture of biopsies taken systematically from the antrum and the corpus. Antimicrobial susceptibility testing to AMO, CLA, and MET was performed. Children having received PPI, H2-blockers, or antibiotics during the 4 weeks preceding endoscopy were excluded. For all of the enrolled children, parental informed consent and child assent was obtained as requested by local ethics policy in the different European countries. The ethics committee of Queen Fabiola Children's University Hospital, Brussels, Belgium, approved the protocol for anonymous data collection and analysis.
The study centers provided the following items on a standardized questionnaire: age, sex, country of birth of the child and the child's mother, previous H pylori infection treatment, concomitant medications, chronic diseases, clinical signs and symptoms at time of endoscopy, and description of endoscopic lesions. Mother's country of birth was used to determine the children's ethnic origin.
During the upper gastrointestinal (GI) endoscopy, biopsies were performed both in the gastric antrum (n = 3) and corpus (n = 3). Two specimens of the antrum and 2 of the corpus were formalin fixed and paraffin embedded before hematoxylin/eosin and Giemsa staining for histopathological analysis according to the updated Sydney classification (19). One antral and 1 corpus biopsy were used for bacteriological culture and antimicrobial susceptibility test using the epsilometer (E) test as previously described (16). Minimal inhibitory concentration breakpoints for resistance were defined as follows: MET ≥8 μg/mL, CLA ≥1.0 μg/mL, and AMO ≥1 μg/mL. These tests were performed locally according to the Clinical Laboratory Standard Institute (formerly National Committee for Clinical Laboratory Standards) recommendations (20). Strict adherence to the methods was mandated for all of the included microbiological laboratories. A previous study showed that E test is a suitable alternative to the agar dilution method (21).
Patients were randomly assigned to receive either a 10-day sequential treatment or a 7-day triple therapy containing OME, AMO, and a third drug (either CLA or MET) prescribed according to antimicrobial susceptibility testing.
This treatment scheme involves an initial 5-day therapy with a combination of OME (10 mg bid below 30 kg body weight or 20 mg bid above 30 kg) and AMO 25 mg/kg bid—maximum 2 g/day), followed by 5 days of OME, CLA (7.5 mg/kg bid—maximum 1 g/day), and MET (10 mg/kg bid—maximum 1.5 g/day).
Triple Therapy According to Antimicrobial Susceptibility Testing
This treatment scheme is a tailored 7-day treatment comprising OME and AMO with CLA in cases of H pylori strains susceptible to CLA or with MET in cases of H pylori strains susceptible to MET and resistant to CLA.
H pylori eradication was assessed by the 13C-urea breath test (UBT) at least 8 weeks after completion of the treatment (22). Briefly, after a fasting period of at least 4 hours, 75 mg 13C-labeled urea was ingested either in a capsule or diluted in 50 mL of water. Then the mouth was rinsed with apple juice, which also served as test meal (a total of 200 mL apple juice was ingested or 100 mL for children younger than 3 years). Six breath samples, 2 each time, were taken at baseline and at 15 and 30 minutes. Ratios of 13C/12C, expressed as Δ 13C in pro mil compared with PeeDee Belemnite calcium carbonate standard, were measured in a stable isotope ratio mass spectrometer. Analytical uncertainties are on the order of 0.1‰. H pylori eradication was considered successful when the delta over the baseline (DOB) remained below 5‰; however, values falling into a gray area between 3.5‰ and 5‰ were controlled after 1 or 2 months. Compliance with treatment was determined by conducting an interview and counting remaining pills and defined as consuming >90% of the medications. Adverse events were assessed by interview plus a specific questionnaire in all of the centers.
For the descriptive analysis of quantitative parameters with calculation of medians and range, the software of StatView (Abacus, Berkeley, CA) was used. The differences between eradication rates were analyzed by the χ2 test. Analysis of variance also was used for the crossing of qualitative and quantitative parameters. The eradication rates with their 95% confidence intervals (CI) and the odds ratios (ORs) were calculated. A P value <0.05 was considered to be significant.
In the intention-to-treat (ITT) eradication rate, all of the patients randomly assigned to one of the treatments were analyzed together, regardless of whether they returned for the posttreatment UBT. In the per-protocol (PP) eradication rate, only patients who performed the posttreatment UBT were considered in the analysis.
Population Characteristics and Endoscopic Findings
During the study period, 165 consecutive children (95 girls and 70 boys; median age 10.4 years, range 2.7–17) undergoing upper GI endoscopy and presenting with H pylori infection were enrolled. Of these 165 enrolled children, 109 were included by the Belgian center, 28 by the French center, and 28 by the Italian center.
Eighty-nine children were born in northern Europe, 20 in southern Europe, 5 in eastern Europe, 26 in northern Africa, 10 in sub-Saharan Africa, 4 in the Middle East, 3 in Asia, and not determined in 8. Their ethnic backgrounds were as follows: northern Europe, 11; southern Europe, 20; eastern Europe, 5; northern Africa, 91; sub-Saharan Africa, 10; Middle East, 4; Asia, 4; and not determined, 20 children.
Median weight at prescription was 37.8 kg (range 15–87) and height 1.43 m (range 0.87–1.82). A previous unsuccessful H pylori eradication treatment had been given to 15 children. Chronic relevant diseases were present in 56 patients (>1 in some children): insulin-dependent diabetes mellitus in 11, allergic manifestations in 7, lactose intolerance in 4, celiac disease in 3, cerebral palsy in 2, epilepsy in 2, recurrent urinary infection in 2, chronic lung disease in 2, autism in 1, extrahepatic biliary atresia in 1, chronic hepatitis B in 1, Glucose-6-phosphate dehydrogenase deficiency in 1, and eosinophilic esophagitis in 1 patient. Concomitant medications were given in 44 patients (>1 in some children): insulin in 11, laxatives in 7, domperidone in 5, nonsteroidal anti-inflammatory drugs in 4, polysaccharide ferric complex in 3, probiotics in 3, salbutamol in 3, antispasmodic in 2, loperamide in 1, ursodeoxycholic acid in 1, rifampicine in 1, and cetirizin in 1 patient.
There were no significant differences between the 2 treatment groups in sex (male/female ratio) or age (median) (1/1.24 vs 1/1.48, 10.0 vs 11.1 years, respectively; P > 0.05). The median weight and height were similar in both groups (33.3 vs 45 kg and 1.38 vs 1.74 m, respectively; P > 0.05). Additionally, the indications for endoscopy were similar in both groups (abdominal pain 68.6% vs 78.0%, nausea/vomiting 13.2% vs 19.5%, heartburn 18.0% vs 14.6%, anemia 8.3% vs 6.0%, and others 14.4% vs 11.0%, respectively; P > 0.05). The other clinical characteristics and endoscopic findings are shown in Table 1.
Distribution of age, sex, indications for endoscopy, previous H pylori eradication treatment, endoscopic lesions, and chronic diseases were similar for the 3 centers except for 11 children with diabetes mellitus type 1 present only in the Belgian center.
Of the 165 children, 15 patients were lost to follow-up, and information about their treatment regimen compliance, adverse effects, or eradication rate was not available and therefore was not included in the PP analysis.
Antimicrobial Susceptibility Testing
Primary resistance to CLA (CLAr) occurred in 24 of 150 (16%) isolated strains and to MET (METr) in 29 of 150 (20%). The antimicrobial susceptibility testing failed in 4 of 150 (2%, H pylori was cultured, but subculture for the E test was not possible for antimicrobial susceptibility testing). Primary resistance rates differ between centers (Italy: CLAr 10/24 and METr 4/24, France: CLAr 5/26 and METr 5/26, Belgium: CLAr 9/100 and METr 20/100). Primary CLAr was significantly more frequent in Italian than in Belgian children (OR 7.2, 95% CI 2.5–20.9, P = 0.0003).
In the 15 children who had previously received an eradication therapy, secondary resistance to CLA occurred in 3 of 15 (20%) children and to MET in 2 of 15 (13%) children.
Eradication was achieved in 127 of 150 (84.6%) children who returned for a follow-up test. The ITT (n = 165) and PP (n = 150) eradication rates are shown in Table 2. Eradication rates were higher using the sequential treatment compared with the tailored triple therapy, but differences were not significant except for ITT analysis with strains susceptible to both CLA and MET (Table 2). The ITT and the PP eradication rates were significantly lower with the sequential treatment in cases of CLA-resistant strains compared with CLA-susceptible strains (Table 2). On the contrary, eradication rates were similar in children after sequential treatment with MET-resistant strains compared with MET-susceptible strains (Table 2). Eradication rates did not differ significantly among the 3 centers.
Adverse Events and Compliance
Abdominal pain was found in 20% of patients (sequential 24%, triple therapy 17%, ns), diarrhea in 14% (12% and 16%, respectively, ns), nausea in 6% (8% and 5%, respectively, ns), and vomiting in 2% (4% and 0%, respectively). One patient receiving triple therapy stopped his treatment prematurely. Except for this patient, all of the patients who returned for the control UBT fulfilled the compliance criteria.
The present pediatric prospective, open-label, multicenter randomized study compared a sequential treatment regimen to a triple therapy tailored to the antimicrobial susceptibility; the eradication of H pylori was higher with the sequential treatment; however, the difference was not significant except for ITT in susceptible strains to both CLA and MET. In the group of children treated with the sequential regimen, the ITT and the PP eradication rates were significantly inferior in case of CLA-resistant compared with CLA-susceptible strains.
In previous studies (17,18), eradication rates >85% and even 90% were observed with triple therapy tailored to the antimicrobial susceptibility in contrast with poorer results in the present study. This lower efficacy of the tailored triple therapy remains unexplained. Although poor or insufficient patient compliance often is responsible for unsuccessful treatment, this was not the case in our series, because noncompliant patients were excluded. Different eradication rates have been reported in different regions of the world. This can be because of not only different host factors but also differences in the characteristics of the strains infecting the different ethnic groups. The 3 centers involved in the present study recruited children living in western European countries but born of migrant families, mainly from northern Africa. This can be an explanation for the lower eradication rates. Antimicrobial resistance acquired during the treatment may be another explanation. Therefore, the comparison of results obtained in different parts of the world need to be critically analyzed with caution, stressing the need for correctly randomized trials. Our results, however, remain consistent with our previous data (4,23) and with the decreasing efficacy of the standard triple therapy in most studies (1).
DOB values obtained in the eradicated group were low (median 0.6‰, range 0.0‰–3.5‰), with only 5 values between 2.0‰ and 2.5‰ and 2 between 2.5‰ and 3.5‰. On the contrary, DOB values obtained in the eradication failure group were high (median 14.6‰, range 5.0‰–77.2‰), with only 3 values between 5.0‰ and 7.5‰, the other values being >8.8‰. The possibility of false-positive UBT results in young children is well documented in the literature (22). This is not the case in our series, because the children with a DOB close to the positive criteria are older than 6 years (9.3–11.5 years).
Triple therapies are the most frequently used treatment in clinical practice in adults and children. A major decline in H pylori eradication rate with these therapies has been observed in the last few years. The sequential regimen is a novel, promising therapeutic approach. A pooled-data analysis of all of the studies on the sequential regimen was performed (24). The eradication rate was calculated, taking into account several factors such as the presence of peptic ulcer disease, the PPI used, the antibiotic susceptibility, and the patient's age including children. Overall, >1800 patients have been treated with the sequential regimen, which was found to be superior to 7- or 10-day triple therapies in adults but also in children and elderly patients, with eradication rates constantly >90% in ITT. Similar results were reported by Jafri et al (25) after analyzing data reported on 4110 adults, in which 1363 were treated by sequential therapy. They reported an eradication rate of 93.4%, significantly superior to the 76.9% eradication rate of triple therapy performed in 2747 patients.
The meta-analysis of Gatta et al (26) evaluated the efficacy of sequential treatment in adults and children compared with triple therapy. In 13 randomized controlled trials, enrolling 3006 adults, the OR for eradication of H pylori with sequential treatment compared with standard triple therapy was 2.99 (95% CI 2.47–3.62) in favor of the sequential treatment. In patients with CLA resistance, the OR for eradication with sequential treatment compared with CLA containing triple therapy was 10.21 (95% CI 3.01–34.58). No difference was found between regimens including tinidazole or MET in the sequential arm. They also analyzed 3 randomized controlled trials enrolling 260 children and adolescents, in which the OR for eradication was (1.98, 95% CI 0.96–4.07) in favor of the sequential treatment, but difference failed to reach statistical significance. No difference in the frequency of adverse effects was found. Our study shows similar results. Gatta et al conclude that the sequential therapy is promising, but further trials are needed in different European countries and in North America before it can be recommended as a first-line treatment.
In these studies, triple therapies are prescribed without tailoring them to the resistance pattern of the strains, which had the effect of markedly improving their efficiencies. In our study, we showed in a prospective, open-label, multicenter study carried out in children from Belgium, France, and Italy that the 10-day sequential therapy regimen is more effective but not significantly better than the tailored triple therapy. Furthermore, the sequential treatment proved to be less efficacious in CLA-resistant compared with CLA-susceptible strains and this could have a significant effect in populations with high CLA resistance rates. It is frequently said that pretreatment antimicrobial testing is expensive and difficult to achieve. Yet, the price of a culture in Belgium is <20 € (US$30), whereas the cost of a treatment is around 70 € (US$100) and the endoscopy with deep sedation approximately 600 € (US$830). Additionally, new techniques such as the fluorescence in situ hybridization (FISH) (27) test can be used when the culture is technically impossible to achieve in a center. The E test used to determine antimicrobial susceptibility in the present study is not accepted worldwide as a valid test, but the susceptibility tests were compared with the FISH technique in the Italian and the Belgian centers and with agar dilution in the Belgian center (data not shown).
Recently, Francavilla et al (13) published a monocentric retrospective study showing that only the A2143G mutation, involved in the resistance of H pylori to CLA, is associated with a higher risk of treatment failure. They concluded that sequential therapy is more effective than classic therapy in case of CLA resistance, but again their triple therapy was not tailored to antibiotic susceptibility. In our study, based on a prospective determination of the antimicrobial susceptibility, we showed, not surprisingly, that the replacement of CLA by MET in CLA-resistant strains resulted in better results with the adapted triple therapy than with the sequential therapy. Therefore, we believe that the sequential treatment should be used as a first-line therapy only when antimicrobial susceptibility testing is available for the patient to treat or in a population in which CLA resistance rate is known to be low, that is, <20%. The latter that takes hypothesis needs validation in an international study into consideration the effect of populations migrating from counties in which CLA resistance rate is high (eg, southern Europe) or from continents or regions with poorly determined CLA resistance rates (eg, Africa, the Middle East) to regions with a low CLA resistance rate (eg, northern Europe).
In conclusion, our prospective, open-label, multicenter study shows that sequential treatment is greatly effective for eradicating H pylori in children, with a higher (although not significantly) eradication rate compared with triple therapy tailored to antimicrobial susceptibility. The eradication rate is significantly inferior with this sequential treatment in children with CLA-resistant strains as compared with CLA-susceptible strains or CLA-MET susceptible ones. The sequential regimen would be useful for a first-line option when H pylori antimicrobial susceptibility testing is not available only in populations with low CLA resistance rates.
1. Vakil N. H pylori treatment: new wine in old bottles? Am J Gastroenterol 2009; 104:26–30.
2. Gottrand F, Kalach N, Spyckerelle C, et al. Omeprazole combined with amoxicillin and clarithromycin in the eradication of Helicobacter pylori in children with gastritis: a prospective randomized double-blind trial. J Pediatr 2001; 139:664–668.
3. Cadranel S, Bontems P, Van Biervliet S, et al. Improvement of the eradication rate of Helicobacter pylori gastritis in children is by adjunction of omeprazole to a dual antibiotherapy. Acta Paediatr 2007; 96:82–86.
4. Oderda G, Shcherbakov P, Bontems P, et al. Results from the Pediatric European Register for Treatment of Helicobacter pylori (PERTH). Helicobacter 2007; 12:150–156.
5. Zullo A, Vaira D, Vakil N, et al. High eradication rates of Helicobacter pylori with a new sequential treatment. Aliment Pharmacol Ther 2003; 17:719–726.
6. Hassan C, De Francesco V, Zullo A, et al. Sequential treatment for Helicobacter pylori eradication in duodenal ulcer patients: improving the cost of pharmacotherapy. Aliment Pharmacol Ther 2003; 18:641–646.
7. De Francesco V, Zullo A, Margiotta M, et al. Sequential treatment for Helicobacter pylori does not share the risk factors of triple therapy failure. Aliment Pharmacol Ther 2004; 19:407–414.
8. Zullo A, Gatta L, De Francesco V, et al. High rate of Helicobacter pylori eradication with sequential therapy in elderly patients with peptic ulcer: a prospective controlled study. Aliment Pharmacol Ther 2005; 21:1419–1424.
9. Francavilla R, Lionetti E, Castellaneta SP, et al. Improved efficacy of 10-day sequential treatment for Helicobacter pylori eradication in children: a randomized trial. Gastroenterology 2005; 129:1414–1419.
10. Lionetti E, Miniello VL, Castellaneta SP, et al. Lactobacillus reuteri therapy to reduce side-effects during anti-Helicobacter pylori treatment in children: a randomized placebo controlled trial. Aliment Pharmacol Ther 2006; 24:1461–1468.
11. Kalach N, Serhal L, Bergeret M, et al. Sequential therapy regimen for Helicobacter pylori infection in children. Arch Pediatr 2008; 15:200–201.
12. Francavilla R, Lionetti E, Cavallo L. Sequential treatment for Helicobacter pylori eradication in children. Gut 2008; 57:1178.
13. Francavilla R, Lionetti E, Castellaneta S, et al. Clarithromycin-resistant genotypes and eradication of Helicobacter pylori. J Pediatr 2010; 157:228–232.
14. Fuccio L, Eusebi LH, Bazzoli F. Can sequential therapy for Helicobacter pylori eradication overcome clarithromycin resistance?: Data still do not support conclusions. Am J Gastroenterol 2010; 105:1674–1675.
15. Tong JL, Ran ZH, Shen J, et al. Sequential therapy vs. standard triple therapies for Helicobacter pylori infection: a meta-analysis. J Clin Pharm Ther 2009; 34:41–53.
16. Koletzko S, Richy F, Bontems P, et al. Prospective multicentre study on antibiotic resistance of Helicobacter pylori strains obtained from children living in Europe. Gut 2006; 55:1711–1716.
17. Arenz T, Antos D, Russmann H, et al. Esomeprazole-based 1-week triple therapy directed by susceptibility testing for eradication of Helicobacter pylori infection in children. J Pediatr Gastroenterol Nutr 2006; 43:180–184.
18. Kato S, Konno M, Maisawa S, et al. Results of triple eradication therapy in Japanese children: a retrospective multicenter study. J Gastroenterol 2004; 39:838–843.
19. Dixon MF, Genta RM, Yardley JH, et al. Classification and grading of gastritis. The updated Sydney System. International Workshop on the Histopathology of Gastritis, Houston 1994. Am J Surg Pathol 1996; 20:1161–1181.
20. National Committee for Clinical Laboratory Standards. Performance standards for antimicrobial susceptibility testing and approved standard M7-A5. Informational supplement M100-S10. Wayne, PA: NCCLS; 2000.
21. Best LM, Haldane DJ, Keelan M, et al. Multilaboratory comparison of proficiencies in susceptibility testing of Helicobacter pylori and correlation between agar dilution and E test methods. Antimicrob Agents Chemother 2003; 47:3138–3144.
22. Kindermann A, Demmelmair H, Koletzko B, et al. Influence of age on 13C-urea breath test results in children. J Pediatr Gastroenterol Nutr 2000; 30:85–91.
23. Bontems P, Devaster JM, Corvaglia L, et al. Twelve year observation of primary and secondary antibiotic-resistant Helicobacter pylori strains in children. Pediatr Infect Dis J 2001; 20:1033–1038.
24. Zullo A, De Francesco V, Hassan C, et al. The sequential therapy regimen for Helicobacter pylori eradication: a pooled-data analysis. Gut 2007; 56:1353–1357.
25. Jafri NS, Hornung CA, Howden CW. Meta-analysis: sequential therapy appears superior to standard therapy for Helicobacter pylori infection in patients naive to treatment. Ann Intern Med 2008; 148:923–931.
26. Gatta L, Vakil N, Leandro G, et al. Sequential therapy or triple therapy for Helicobacter pylori infection: systematic review and meta-analysis of randomized controlled trials in adults and children. Am J Gastroenterol 2009; 104:3069–3079.
27. Caristo E, Parola A, Rapa A, et al. Clarithromycin resistance of Helicobacter pylori strains isolated from children’ gastric antrum and fundus as assessed by fluorescent in-situ hybridization and culture on four-sector agar plates. Helicobacter 2008; 13:557–563.