Background:This is a report of the results of a multicenter study performed in children with dyspepsia from five pediatric centers in Puglia, a region in southern Italy. In the study, clinical features of Helicobacter pylori infection, the reliability of diagnostic techniques, and the involvement of bacterial strains were examined.Methods:Fifty‐three outpatients with dyspepsia enrolled in our study and compiled a diary recording clinical symptoms in patients before they underwent the following diagnostic techniques: endoscopy, biopsy for histologic analysis, rapid urease test, 13C urea breath test, serology specific for immunoglobulin (Ig) G and anti‐CagA and VacA.Results:H. pylori showed a prevalence of 30.2% (n = 16). Histologic positivity was seen in all patients at the antral level (H. pylori‐associated chronic gastritis). In the gastric body, bacterial chronic active gastritis was present only in six patients (H. pylori‐associated chronic pangastritis). Clinical evaluation showed a significant difference in favor of subjects positive for H. pylori only for epigastric burning and/or pain (p < 0.001). The comparison of results of diagnostic tests, using histology as the gold standard, showed sensitivity and specificity of more than 93% for 13C urea breath test and more than 85% for rapid urease test and serology. Anti‐CagA antibodies were found in 64.3% and anti‐VacA antibodies in 42.8% of H. pylori‐positive patients.Conclusions:H. pylori prevalence in children with dyspepsia from the geographic area studied in comparable with that found in other developed countries. Approximately 50% of the studied patients were infected by cytotoxic strains. The urea breath test was the most reliable noninvasive diagnostic tool and is suitable for routine use, although endoscopy with histologic assessment remains the definitive investigation and is particularly important in patients with positive serology for CagA and VacA. Finally, the frequency of aggressive strains in our region seems to affect the clinical pattern; this emphasizes the importance of definitive diagnosis in children and offers a new role for serology.