Study design and setting
The study was designed to conduct into two consequential phases as detailed hereafter:
Phase I: a cross-sectional design was carried out in the hematology clinic and inpatient of the pediatrics department during the period from January to December 2019 to explore prevalence of H. pylori among chronic ITP children.
Phase II: interventional design among chronic ITP children who were reported positively for H. pylori to investigate the impact of treatment of H. pylori infection on platelet count response.
The study was conducted on one hundred children, through their regular visits to the hematology clinic for follow-up. Target children aged 02–18 years old and diagnosed with chronic ITP through convenient sampling.
Based on the criteria of the American Society of Hematology (ASH) , children were diagnosed with ITP when the initial platelet count is < 100000 to avoid the confusing effect of the incident ITP therapies. Eligible children are those who have platelet count above 20000 and below 100000.
Children with other causes of thrombocytopenia such as HCV, HBV, or HIV, drugs, lymphoproliferative disorders, other auto-immune disorders, and children with active life-threating bleeding at the time of recruitment were excluded.
The number of participants was estimated using the Epi-Info version 7 StatCalc designed by the Centers for Disease Control and Prevention (CDC) and the World Health Organization (WHO). We exceeded the least required sample size to enhance statistical power and avoid unpredicted low response rates.
The authors filled out a comprehensive questionnaire for each patient, involving detailed history and physical examination. We collected a venous blood sample from each patient for laboratory investigation, including CBC with a direct platelet count. Finally, stool samples were collected in clean containers used for the detection of H. pylori Ag; colored chromatographic immunoassay for the qualitative detection of Helicobacter pylori antigen using CerTest H. pylori detection kit (CERTEST BIOTEC, Spain). It is a strip consisting of a nitrocellulose membrane pre-coated with mouse monoclonal antibodies against H. pylori on the test line (T) and with a rabbit polyclonal antibodies against a specific protein on the control line (C).
Standard H. pylori eradication therapy
Treatment of H. pylori infection in the form of amoxicillin 50 mg/kg day, clarithromycin 15 mg/kg/day for 2 weeks, and proton pump inhibitor 1 mg/kg/day was given to all cases for 1 month.
Reevaluation of their H. pylori status and platelet count was done 6 months after stopping the treatment to assess the efficacy of treatment of Helicobacter infection (whether the patient was cured of H. pylori infection or not) and to investigate the effect of H. pylori eradication on the patient’s platelet count.
The study was approved by the Research Ethics Committee of the Faculty of Medicine. The authors explained the aim and procedures of the study for parents and assured the confidentiality of the data collected, as well as that of the laboratory tests. We got informed consent from parents of children prior to the recruitment.
We applied statistical analysis using SPSS version 15 (Statistical Package for the Social Science; SPSS Inc., Chicago, IL, USA). Data were presented as frequencies, percentages, and mean ± standard deviation when appropriate. For numerical variables, the Mann-Whitney U test or Kruskal-Wallis test was used for independent samples and Wilcoxon signed-rank test for paired (matched) samples. For categorical data, the chi-square (χ2) test was performed. In case of expected frequency of less than 5, the exact test was applied instead. Spearman rank correlation equation was used for the correlation between various variables. P values less than 0.05 were considered statistically significant.