Moroccan Children With Helicobacter pylori Infection: Demographics, Clinical Features, and Histological Findings

Background: Infesting nearly half of the world's population, Helicobacter pylori is thought to cause peptic ulcers and gastric adenocarcinoma. Several studies have examined the association between H. pylori and socioeconomic, clinical, and histological factors in pediatric populations. Similarly, this study aimed to describe the characteristics of H. pylori infection in Moroccan children. Methods: Patients aged 1-17 years who underwent upper gastrointestinal endoscopy over a period of two years from January 2019 to January 2021 were included in this study. Gastric biopsies from the antrum and corpus of the stomach were collected. Detection of H. pylori infection was confirmed by Giemsa stain. Demographic data and clinical and endoscopic characteristics were collected and histopathological findings with gastritis scoring were recorded according to the Sydney System. Results: In 213 children, 95 (45%) were found to be infected with H. pylori, and the infection rates increased as the children aged. While no significant relationship between the infection of H. pylori and all symptoms was founded, a significant association was found in nodular gastritis (p<0.05), and 98% of the infected children had chronic inflammation, which was active in 22% and atrophic in 47%. The atrophy and activity were absent or mild, and the inflammation was mild to moderate. Conclusion: According to this study, nodular gastritis and nonspecific symptoms were related to H. pylori infection in Moroccan children. In addition, the association between this disease and gastric atrophy in our study needs the monitoring of the mucosa of Moroccan children with gastritis and identifying factors that may contribute to gastric cancer.


Introduction
Helicobacter pylori is one of the most prevalent human pathogens and the key pathogenic bacterium in pediatric gastroenterology. Affecting more than half of the world's population, this bacterium continues to be a major public health issue [1]. It is a major cause of chronic gastritis, and peptic ulcer diseases, and a significant risk factor for stomach cancer and mucosa-associated lymphoid tissue (MALT) lymphoma developed in adults [1].
A published study on the pediatric population found that the prevalence rates of H. pylori infection range from 1.8% to 65% among children [2]. However, data on the prevalence of this infection in Morocco is limited to adults, such as the 69.4% detected by biopsy culture in Rabat [3]. The diagnosis of H. pylori infection is possible by invasive and non-invasive tests. Among the invasive methods, histology was the first method used to detect H. pylori providing additional and essential information on the status of the mucosa [4]. Much is known about the association of H. pylori infection with gastrointestinal symptoms in adults, such as nausea, vomiting, and abdominal pain [5], but its role in children is unclear. The clinical manifestations of this infection at this age are often modest, and nonspecific [6]. In addition to symptoms as clinical factors of H. pylori infection, factors such as age, gender, and a variety of socioeconomic indicators are associated with the prevalence of H. pylori infection [7].
In addition, another aspect of H. pylori infection is its histopathological characteristics. As a pathogen, H. pylori is highly adapted to the gastric mucosa, which causes gastritis in all infected individuals. Most pediatric studies report histological gastritis in all H. pylori infections . The proportion of patients with this   1, 2  1, 2  1, 2  1, 2  1,   3  1, 3  4, 5  4, 5 inflammation ranges from 75% to 100% [8]. In addition to peptic ulcers and gastric cancers, H. pylori gastritis has been described by several studies, which reported the presence of H. pylori with the histological lesions of gastric mucosa in the pediatric population [9]. Accordingly, this study examined and described demographical, clinical, endoscopic, and histopathological aspects of H. pylori infection in Moroccan children. To the best of our knowledge, this is the first study of its kind in our context.

Materials And Methods
A cross-sectional study was conducted at the Hassan II University Hospital in Fez, Morocco, from January 2019 to January 2021. Outpatients (under 17 years of age) who required diagnostic or therapeutic upper gastrointestinal endoscopy and had not taken a drug within 30 days of the testing (antibiotics or proton pump inhibitors) were included randomly in this study. Informed consent and a structured questionnaire containing sociodemographic and clinical information were obtained from the patient's parents as approved by the Research Ethics Committee of the University Hospital of Fez, Morocco (approval number: 28/20).
The endoscopic lesions were recorded and biopsy specimens were captured during endoscopy supervised by a medical gastroenterologist with the support of trained nurses and auxiliary staff. Two biopsy specimens were collected, one at the antrum and one on the corpus. They were fixed in 10% formalin, embedded in paraffin, and cut at 6 mm. Histological sections were stained with hematoxylin and eosin (H&E) and modified Giemsa for light microscopy [10]. They were analyzed according to the revised Sydney System [11] by a pathologist who noted the presence or absence of H. pylori and the histopathological funding. The histological variables (topographic of gastritis inflammation, intensity of inflammation, gastritis activity, gastric atrophy, intestinal metaplasia, and intestinal dysplasia) were graded according to the updated Sydney System visual analog scale to generate a score (0 = absent, 1 = mild, 2 = moderate and 3 = severe). Subjects were considered infected when the H. pylori bacterium was detected in histological sections stained with Giemsa.
The mean age and the proportions of the categorical data were calculated using a simple descriptive analysis.
Bivariate analysis was carried out using the chi-square (χ 2 ) test and the binary linguistic for comparing and exploring the distribution of the categorical variables in the infected and no infected children groups. A pvalue ≤ 0.05 was considered significant.

Results
In this study, 213 children with symptoms (age range 1-17 years) were included of which 116 (54%) were females and 97 (46%) were males. According to the results of the Giemsa stain ( Figure 1), 95 children (45%) presented H. pylori infection and 118 (55%) did not ( Table 1). Among several factors studied about H. pylori positivity, the factors tested in the present study were gender and age. Table 2 shows the frequency of H. pylori concerning gender and age group. There was no statistical difference in H. pylori frequency in gender ( Table 2). Out of 97 males, H. pylori was positive in 47%, and of 116 females, it was positive in 44%. There was, however, a significant association between H. pylori infection and age ( Table 2). The rate of a positive number increased with children's age ( Table 2) from 26% in the age group of one to four years to 65% in the older age groups (12-17 years). In the age group of 5-8 years and 9-12 years, we recorded rates of 49% and 52%, respectively.

FIGURE 1: Helicobacter pylori in Giemsa staining
A: Helicobacter pylori bacterium   As shown in Table 3, vomiting in 79 cases (37%) followed by epigastric pain in 47 patients (22%) were the most common clinical indications for endoscopy. There was no significant relationship between H. pylori infection and all of the symptoms cited in this study. According to the gastric endoscopy findings (

TABLE 4: Endoscopic findings in all enrolled patients
On histopathological examination of 213 specimens (Table 5), the lesions of chronic inflammation observed in a majority of the infected children (98%) were atrophic in 91 cases (47%), active in 42 patients (22%), and with lymphoid follicles aspect in 56 cases (57%). Among the infected group, and according to the gastritis grading, the chronic inflammation intensity ranges from mild in 83% to moderate in 11% and severe in 4%; the activity of gastritis was mainly absent in 72% or mild in 26% and the atrophy varied from mild (48%) to moderate (5%).

Discussion
H. pylori was detected in the present study by histological examination based on Giemsa staining. This is a method of choice, cheap, easy to perform, and reproducible. It is significantly more sensitive than other special stains as H&E used to appreciate the gastric mucosa [12].
We found that the prevalence of H. pylori infection in our study (45%) was similar to that in a study conducted on symptomatic Saudi children (49%) [13], and it was higher than 24.7%, and 14.6% reported respectively in a Brazilian pediatric and adolescent population [14], and in Jordan [15]. Thus, our result confirms a higher prevalence of H. pylori in the Moroccan population just like a study conducted in our area in a group of 429 adults (69.9%) [16]. The Rabat region reported a rate of 69.4% based on the results of a multicenter study on symptomatic adults [3].
The higher prevalence of H. pylori infection could be attributed not only to the sociodemographic, environmental, behavioral, and clinical factors but also to other factors that could significantly influence the variation in results among the studies, including inclusion criteria, sample size, method of diagnosis, and presence of the gastrointestinal symptoms, which provided a growth medium (changing pH, thinned gastric wall, gastric ulceration, and altered gut microbiota) for the bacteria, so the participants were more likely to contract H. pylori [17].
Among demographic factors, there are no differences in H. pylori infection rate in both girls and boys (p=0.513, OR = 1,198) in our study. Regarding age, similar to that reported by other authors [18], the prevalence of this infection increased with age and reported values of 26%, 49%, 52%, and 65% in the age groups of one to four years, five to eight years, 9-11 years, and 12-17 years, respectively. In addition, there was an increased rate of H. pylori infection in older children, which could be explained by person-to-person transmission of the bacterium. These factors are restricted to family members during neonatal and preschool years, reducing the risk of H. pylori gastroenteritis in the youngest children compared with the older school-age children exposed to more sources of infection in an environment [19]. In this direction, it is reported that the low infection rate of preschool children (0.6%) was significantly increased in high school students (13.5%) (P =0.001) [19].
Regarding the clinical factors of H. pylori infection, the population evaluated in this study was characterized by symptoms represented mainly in celiac disease suspicion, dysphagia, gastroesophageal reflux disease, vomiting, and epigastric pain. The current study showed that the children's symptoms distribution for H. pylori-positive cases was not statistically significant and similar to that reported by Altamimi et al. who reported no association between the majority of gastrointestinal symptoms and being infected with H. pylori [15]. In addition, according to some authors, the H. pylori infection was associated with epigastric pain. But it was mainly either clinically silent or associated with nonspecific signs or symptoms according to results of various childhood studies [20].
In endoscopy, among the infected group, the current study showed that nodular gastritis was the most common gastritis status related to H. pylori infection. It was seen in 42 cases (64.61%), higher than that reported by Luzza et al. (40%) [21]. Similar to our results, and according to several authors, nodular gastritis was the only which showed a significant statistical association with H. pylori infection and may be a pathognomonic macroscopic finding of childhood H. pylori infection [22]. In this respect, the positive predictive value of this endoscopic status in H. pylori infection ranged between 46% and 73% in the literature [23].
Histologically, our results demonstrated that chronic gastritis was mainly seen in the infected group (98%), similar to that found by Mazigh-Mrad et al. (88.5%) [24]. A similar rate (87.7%) was reported by Al Kirdy et al. in infected patients [25]. The same study showed that active gastritis was seen in most of the infected patients (84.9%). A value of 63% was reported by Mazigh Mrad et al. [24], higher than that shown in our study.
Regarding gastritis degree, chronic gastritis was commonly mild in 83% and moderate in 11% of the infected cases in our study. It was severe in only four infected children (4%). Similarly, Cárdenas-Mondragón et al.
reported that H. pylori infection was usually associated with mild to moderate inflammation in children [26]. In addition, according to the analysis of these authors, the rarest cases of severe gastritis could be explained by coinfection H. pylori bacterium and Epstein Barr virus [26]. Regarding the activity of gastritis and similar to that registered by authors [26], It was mild in 26% to absent in 72% of the infected group in our population.
The rate of gastric atrophy in the infected group was higher (53%) than that reported by several authors in another context as Guiraldes et al. [8], and Campbell et al. [27] with values of 0 and 2/83. It was lower than that reported in a Turkish country (72%) characterized by a similar prevalence of H. pylori to our context (43.9-53%) [28]. In addition, gastric atrophy was not only observed in adult patients [29], it was estimated to vary from 0% to 72% in different studies in pediatric populations [30], which suggests the need for close and extended clinical and endoscopic surveillance of children with atrophic gastritis and identifying other factors (excluding H. pylori) that may predispose them to gastric atrophy [30].
The principal limitation of this study was the exclusion of eradication results and the therapeutic strategy used for managing our patients. It was not possible due to the study design and the inclusion of different diseases of the stomach. Thus, we considered the need for further research using an eradication test for H. pylori infection as a urea breath test and focusing on one of the stomach diseases individually.
In the present study, we were able to characterize H. Pylori infection in Moroccan children, where this infectious disease was characterized by nodular gastritis and gastric atrophy, indicating that clinical, endoscopic, and histopathological findings should be combined for monitoring the mucosa of children.

Additional Information Disclosures
Human subjects: Consent was obtained or waived by all participants in this study. Research Ethics Committee of the University Hospital of Fez, Morocco issued approval 28/20, December 14, 2020. Animal subjects: All authors have confirmed that this study did not involve animal subjects or tissue. Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work.