Michel Bergeret

The 13carbon urea breath test for the noninvasive detection of Helicobacter pylori in children: comparison with culture and determination of minimum analysis requirements.

BACKGROUND The purpose of the study was to determine the accuracy of the labelled 13carbon urea breath test for the diagnosis of Helicobacter pylori in children and to simplify the 13carbon urea breath test in identifying the most discriminating sampling time. METHODS H. pylori was searched for in 100 children aged 10.5+/-4.5 years by histology, bacteriological counts, and culture on antral biopsies together with serology and 13carbon urea breath test. Breath samples were obtained before ingestion (T0) of 75 mg urea-13C and every 10 minutes after until T60. 13CO2 excess ratio was measured by isotope ratio mass spectrometry, and values expressed as delta per mil over baseline enrichment (delta 13CO2). The arithmetic mean (Mdelta 13CO2) of T20 to T60 values was calculated and the test considered positive with Mdelta 3CO2 higher than Mdelta 13CO2 + 3 SD as determined in noninfected children. RESULTS Mdelta 13CO2 of noninfected children as assessed by culture was 1.4+/-0.6 per mil, determining a positive cut-off value of 3.44 per mil. Mdelta 13CO2 was correlated in 11 children with biopsy bacteriological counts. Both culture and 13carbon urea breath test were positive in 38 of 100 children, without any discordance. Plotting 13carbon urea breath test results at each sampling time versus Mdelta 13CO2 showed weaker correlations at T20, T30, T50, and T60, than at T40. The two-sample method at T0 and T30, T40, T50, had high sensitivity and specificity. Single-sample analysis obtained at T40 gave a comparable sensitivity and a slightly reduced specificity. CONCLUSION 13carbon urea breath test is sensitive and specific in children. Two samples collected at T0 and T40 provide the most discriminating procedure.

Genetic and transmission analysis of Helicobacter pylori strains within a family.

Point mutations, intragenic recombination, and introduction of foreign alleles enhanced strain diversity within the family.

High levels of resistance to metronidazole and clarithromycin in Helicobacter pylori strains in children.

ABSTRACT The aim of the study was to evaluate the prevalence of resistance to amoxicillin, metronidazole, and clarithromycin before treatment ofHelicobacter pylori infection in children and to assess the evolution of resistance with time. The study was carried out between 1994 and 1999 with 150 H. pylori-positive children through gastric culture (antimicrobial susceptibility) and histology. All cultured H. pylori strains were sensitive to amoxicillin, 64 (43%) were resistant to metronidazole, 32 (21%) were resistant to clarithromycin, and 14 (9%) were resistant to both metronidazole and clarithromycin. The overall prevalence of resistance to metronidazole and clarithromycin did not change significantly with time. The study highlights the generalized high-level and stable metronidazole and clarithromycin resistance of H. pylori strains from children.

Immunoblotting and serology for diagnosis of Helicobacter pylori infection in children.

Background. The easiest way to identify the presence of current or past Helicobacter pylori infection is to test for antibodies. The aim of this study was to compare an enzyme‐linked immunosorbent assay (ELISA) technique based on the detection of IgG antibodies directed against a global antigenic preparation with immunoblotting based on the analysis of IgG antibody reactivity to separate proteins. Methods. Sera were collected from 80 children (mean age, 9.9 ± 4.3 years). The reference tests were microbiologic and histologic examination of gastric biopsies obtained at upper endoscopy. Results. The immunoblotting was more sensitive (100%) and specific (88%) than ELISA (96 and 79%, respectively) in the evaluation of H. pylori infection in children. Its positive predictive value was 92%, and its negative predictive value was 100%. The best performance index of immunoreactive bands to detect antibodies was obtained with the 26‐kDa (88.7%), 30‐kDa (77.5%) and 19.5‐kDa (70%) antigens. Antibodies by immunoblot technique against the CagA antigen were present in 43.1% of children. Conclusion. Immunoblotting is highly sensitive and more specific than ELISA in children and provides additional information about the full serologic profile. Immunoblotting may therefore be a useful complement to serology, particularly in cases with doubtful ELISA results.

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

Background:  The aim of our study was to assess the different mutations involved in clarithromycin‐resistant Helicobacter pylori strains isolated from French children and their temporal trends.

Using macro-arrays to study routes of infection of Helicobacter pylori in three families.

Background Analysis of the evolutionary dynamics of Helicobacter pylori allowed tracing the spread of infection through populations on different continents but transmission pathways between individual humans have not been clearly described. Materials and Methods To investigate person-to-person transmission, we studied three families each including one child with persistence of symptoms after antibiotic treatment. Ten isolates from the antrum and corpus of stomach of each family member were analyzed both by sequencing of two housekeeping genes and macroarray tests. Results A total of 134 (8.4%) out of the 1590 coding sequences (CDSs) tested, including cag PAI and insertion sequences, were present in some but not all isolates (and are therefore defined as variable CDSs). Most of the variable CDSs encoded proteins of unknown function (76/134) or were selfish DNA including that encoding restriction/modification enzymes (13/134). Isolates colonizing the stomach of one individual can vary by point mutations, as seen in hspA, or by the gain or loss of one to five CDSs. They were considered as (genetic) variants. The phylogenetic clustering of gene profiles obtained on macro-arrays allowed identifying the different strains infecting families. Two to five strains circulated within a family. Identical strains were present in at least two members of all three families supporting the accepted model of intrafamilial transmission. Surprisingly, the mother was not implicated in the transmission of H. pylori in the two French families. Sibling-to-sibling transmission and acquisition of H. pylori from outside the family appeared to be probable in the transmission pathways. Conclusion Macroarray analysis based on previously selected CDSs gives a comprehensive view of the genome diversity of a pathogen. This approach combined with information on the origin of the hspA and glmM alleles revealed that Helicobacter pylori infection may be acquired by more diverse routes than previously expected.

Échees du traitement antibiotique des salmonelloses sévères de l'enfant et utilisation des quinolones

BACKGROUND: Quinolone antibiotics are effective in the treatment of Salmonella infections in adults. Their use in children is limited by their side-effects. POPULATION AND METHODS: Forty-two patients (21 girls and 21 boys), aged 1 month to 12 years (mean 3.3 yrs) were admitted from September 1991 to June 1993 for severe Salmonella infections. Criteria of severity were persistent diarrhea and fever for more than 3 days. Thirty-one of these patients were less than 5 years of age. Blood culture was positive in 7 out of 35 patients: culture of the stools was positive in all patients. Five of the 42 patients had presented an acute episode of Salmonella infection a few weeks earlier and had remained asymptomatic carriers until the new acute and severe episode of diarrhea. All patients were given usual antibiotics, mainly ampicillin, amoxicillin, trimethoprime-sulfamethoxazole. Twenty-five of these patients were then given pefloxacin, 12 mg/kg/day, since the 5th day, for 7 days, because persistence of diarrhea and fever. RESULTS: Diarrhea and fever disappeared within less than 2 days in the group of patients given pefloxacin, even though in 6 patients the infecting Salmonella was in vitro resistant to beta-lactamins. Twenty % of patients remained asymptomatic carriers of Salmonella in the group treated by pefloxacin vs 47% in the group without it. There was no difference in species of Salmonella between both groups. None of the patients treated by pefloxacin developed side-effects during the six months following its administration. CONCLUSIONS: Short treatment by pefloxacin may be an alternative choice for treating severe Salmonella infections in children.

Serum levels of pepsinogen I, pepsinogen II, and gastrin-17 in the course of Helicobacter pylori gastritis in pediatrics.

To the Editors: as a comment to the editorial of Oderda et al: In an interesting editorial in this issue of the journal, Oderda et al (1) review which tests may be used to diagnose Helicobacter pylori (H. pylori) in children. Some are invasive (i.e. upper gastro-intestinal endoscopy and multiple gastric biopsies), and some are non invasive (i.e. serology, C-urea breath test and stool antigen determination). All exhibit various advantages or disadvantages, and for that purpose, their respective place in the diagnosis decision tree differ. In symptomatic children, as clearly stated by the position papers of both the European Task Force for H. pylori infection in children (2) and NASPGAN (3), upper gastrointestinal endoscopy with gastric biopsies remains the gold standard method for the diagnosis of H. pylori gastritis in children. Non-invasive tests are non longer recommended to screen children with abdominal pain. Some new approaches may currently be suggested. Recently, a new EIA blood panel test (Biohit , Helsinki, Finland), has been developed, associating group-I pepsinogens (PGI), group-II pepsinogens (PGII), gastrin-17 (G-17), and H. pylori antibodies, and proposed as a non endoscopic tool for diagnosis and screening of chronic and atrophic gastritis in adults (4–6). In children, such an alternative procedure has not yet been investigated. PGI are synthesized solely in the oxyntic glands and mucus neck cells of the gastric corpus, whereas, PGII are uniformly formed in the glands of the entire stomach and to some extent in the Brunner glands in the first part of the duodenum. In case of atrophic corpus gastritis, the level of serum PGI decreases, whereas the level of PGII remains stable or decreases slightly. Thus, the level of the serum PGI/PGII ratio decreases proportionally to the severity of corpus atrophic gastritis (4). Gastrin is synthesized in G-cells, which are found in the antrum, the most important one being G-17. In case of atrophic antral gastritis and loss of antral G cells, serum G-17 remains low although the stomach is achlorhydric or hypochlorhydric (4). In this new EIA blood panel test, panels include assays of postprandial (after stimulation with protein-rich drink) serum gastrin (G-17prand) which seems to be much more accurate with a higher sensitivity and specificity as compared to basal fasting serum gastrin (G-17fast) (5,6). Using an algorithm, this blood test panel is able to establish with high sensitivity and specificity whether the patient has gastritis, whether this gastritis is atrophic or not and in which part of the stomach the atrophic changes are located. The algorithm (decision tree) is used for classification of patients into different categories of atrophic gastritis by the H. pylori antibody titers, upon the cutoff levels of PGI and G-17prand. The absence of evidence of H. pylori infection is considered to indicate an autoimmune origin of gastritis (4). Finally, H. pylori gastritis, without gastric atrophy, tends to raise the serum levels of G-17 and PGI. A low intragastric acidity increases the serum levels of G-17 and vice versa. Permanent, long-lasting hypoor achlorhydria results in extremely high levels of circulating G-17, possibly due to antral G-cells hyperplasia (6). The aim of our study was a preliminary assessment of this new non endoscopic blood panel test in the course of H. pylori related gastritis in pediatrics. An observational retrospective study was performed in 100 children, mean age 10 years (1–18), 52 females and 48 males, who underwent diagnostic upper-gastrointestinal endoscopy for various dyspeptic symptoms, recurrent abdominal pain and vomiting. Gastric biopsies were taken in antrum and corpus, at least 2 from each site, for histologic and bacterial culture assessment. Histological analysis, H. pylori, chronic inflammation, activity, atrophy, and intestinal metaplasia were noted and graded according to the updated Sydney System. H. pylori infection was considered for gastric biopsies with positive histology and/or culture. Within the same day, fasting serum samples were also analysed using the EIA blood panel tests for H. pylori antibodies (cut-off levels .38 EIU), PGI, PGII and G-17fast (Biohit , Helsinki, Finland). Non parametric MannWhitney U test was used in the calculation of the significance between groups, the StatView Software (Abacus , CA, USA) values were considered significant when P , 0.05. The sensitivity, specificity, positive and negative predictive values and test accuracy of the H. pylori antibodies were calculated and compared to the results of gastric biopsies. Finally, written parental consent was obtained from all. There were 64 H. pylori positives (Hp+ve) and 36 H. pylori negatives (Hp2ve). Among them 61 Hp+ve (95.3%) and 3 Hp2ve (8.3%) had evidence of chronic superficial gastritis, mild (25), moderate (21) and severe (13). None exhibited gastric atrophy or intestinal metaplasia. Serum PGI and PGII levels showed a significant increase and the PGI/PGII ratio significantly decreased in Hp+ve children as compared to Hp2ve ones, P , 0.0002. No significant variation of G-17fast serum level occurred between Hp+ve and Hp2ve children, Table 1. Compared to gastric biopsies, H. pylori antibodies showed 3 false positive and 23 false negative results. Their

Helicobacter pylori in Children: Acquisition of Antimicrobial Resistance after an Initial Course of Treatment

According to a recent consensus statement concerning Helicobacter pylori infection in children (3), upper gastrointestinal endoscopy with biopsies is the preferred method for establishing an etiologic diagnosis of infection. The treatment recently recommended for children combines a gastric acid inhibitor, usually a proton pump inhibitor (PPI), with two antimicrobial agents, an antimicrobial agent plus a bismuth salt, or two antimicrobial agents (4). In France, where the use of bismuth salts is not authorized, treatment consists of amoxicillin plus either clarithromycin or metronidazole. The consensus statement (3) also proposed a strategy for reevaluating those who remain infected after an initial course of therapy: repeat endoscopy with culture and antimicrobial susceptibility testing, with secondary treatment based on susceptibility test results.

Pefloxacin after Failure of Initial Antibiotic Therapy in Children with Severe Invasive Salmonellosis

Auoroquinolones are widely used for the treatment of enteric fever or invasive salmonellosis in adults (Asperilla et al. 1990). However, they are not licensed for use in children, in whom they are administered only for severe infections. Thus, for susceptible strains, the treatment of enteric fever and septicaemic salmonellosis in childhood is generally ampicillin, amoxicillin or cotrimoxazole (Hornick 1987). The persistence of severe clinical symptoms, despite therapy with antibiotics possessing good in vitro activity, raises the question of the real efficacy of the treatment. We report here 7 cases of severe salmonellosis in children with clinical failure of a commonly used initial antibiotic treatment that is active in vitro, followed by a dramatic improvement after short term therapy with pefloxacin. 1. Patients and Methods