Seroprevalence of Anti-Helicobacter pylori and Anti-CagA IgG Antibodies in Iranian Dyspeptic Patients

1Department of Toxicology and Pharmacology, School of Pharmacy, Kerman University of Medical Sciences 2Department of Research and Technology, Kerman branch, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Kerman, Iran 3Gastroenterology and Hepatology Research Center, Institute of Basic and Clinical Physiology Sciences, Kerman University of Medical Sciences, Kerman, Iran 4Pharmaceutics Research Center, Institute of Neuropharmacology, Kerman University of Medical Sciences, Kerman, Iran Int J Enteric Pathog. 2018 August;6(3):56-59


Background
Helicobacter pylori is a fastidious and microaerophilic gram-negative bacterium infecting the epithelial lining of the stomach that was initially isolated from a human gastric biopsy in 1983. 1 Cytotoxin-associated gene A (CagA) is one of the most studied virulence factors of H. pylori.The positive strains for CagA have been more closely associated with gastric disease compared to CagAnegative strains.The CagA toxin is encoded by the cagA gene as part of the pathogenicity island. 2 This gene, which is associated with type I strains, is responsible for severe forms of gastrointestinal diseases and is present in about 50%-70% of H. pylori strains. 3The presence of CagA-positive strains in the human stomach might be associated with gastritis, peptic ulcer, duodenal ulcer, and recently the development of gastric carcinoma. 4,5elicobacter pylori has worldwide distribution, but its prevalence varies among different ethnicities and countries. 6Low socioeconomic status, low level of education, crowded living places in childhood, consumption of alcohol and poor oral hygiene, gender and occupation are among the risk factors for H. pylori infections. 7Several epidemiological studies have shown that a continuous decrease of H. pylori prevalence is associated with an improved standard of living, 8,9 but many epidemiological aspects of this infection still remain unknown. 10This infection is highly prevalent in developing countries, being higher in less developed countries, [11][12][13] and common in 57%-91% of the Iranian population. 10,14In one study conducted in 2013 in Iran including southeast Iran, the H. pylori seroprevalence rate was 77.6% (for patients aged 1 to 90 years). 10Despite its high prevalence in Iran, a representative cross-sectional study on this infection and its CagA status has not been available in this area.
The purpose of the current study was to investigate the H. pylori prevalence and its virulence factor (CagA-toxin) seroprevalence in dyspeptic patients.Two serological kits were used to detect the anti-H.pylori and anti-CagA IgG antibodies.

Samples
For this prospective cross-sectional study, a total of 659 serum samples were collected from Iranian dyspeptic patients in 2016.Individuals' demographics, including sex, age, and medical historical data were collected.They were aged 17 to 72 years, (mean age ± SD = 40.42± 11.32 years), stratified into 10-20, 21-30, 31-40, 41-50, 51-60, and >61 years age groups (Table 1).A total of 91 serum samples were randomly selected from the investigated population to check the anti-CagA IgG status.

Immunoassay Test
The commercial enzyme-linked immunosorbent assay (ELISA) kits for H. pylori IgG (Monobind, USA) and CagA IgG (Euroimmun, Germany) were used to evaluate the presence of anti-H.pylori and anti-CagA IgG antibodies, as described by the manufacturers.The plates were read at 450 nm by an Elisa reader (Tecan, Switzerland), and the seropositive status was determined.

Statistical Analysis
Data entry was performed and statistical significance was determined using the nonparametric Pearson chi-square, Mann-Whitney and Kruskal-Wallis tests in SPSS software, version 21.0 (IBM corporation).The Pearson's chi-square test was used to compare the categorical data and to find the significance in sex and age groups.Logistic regression tests were run to check the relationship between H. pylori prevalence and variables.The P value less than 0.05 was considered statistically significant.

Results
Table 1 shows the number of tested subjects according to the sex and age groups and their anti-H.pylori IgG seroprevalence.A total of 383 patients (58.1%) were positive for this antibody.Males had a seroprevalence of 54.5% compared to 60.3% recorded for females (P = 0.08).General H. pylori IgG prevalence increased progressively with age (Figure 1), and this was higher in the patients aged 51-70 years when compared to the patients with other ages (P < 0.05).
The seroprevalence of anti-CagA IgG for female and male patients were 51.1% (23 of 45) and 58.7% (27 of 46), respectively.The results also showed a seropositivity rate of 52.9% (37 of 70) and 61.9% (13 of 21) among seropositive and seronegative patients for anti-H.pylori IgG, respectively.

Discussion
The overall H. pylori seroprevalence was found to be 58.1% among Iranian dyspeptic patients.This is lower than earlier rate in Iran, which were reported as seropositivity rates of 77.6% in southeast Iran, 10 67.1% in southern Iran, 14 and 86.8% in endoscopy unit of a hospital in Tehran 15 among dyspeptic patients.The detected H. pylori seroprevalence in this study was also lower than the reports from Afghanistan, 16 and Pakistan, 17 but higher than those from other Asian countries, 18,19 European countries (like England, France, Italy, Scandinavia, and Belgium), and the United States. 7,20These seroepidemiological studies were conducted in specific subgroups of the population.
Helicobacter pylori IgG prevalence in Iranian dyspeptic population may be related to socioeconomic and living conditions as risk factors.This may be explained by the association between H. pylori, and low socioeconomic status, low level of education, crowdedness, poor oral hygiene, environmental sanitation, and eating spicy foods. 7,21There are reports on significant risk of H. pylori infections for men, but others disproved this dependency. 22his was approved in the current study (P = 0.08), the same as a previous report. 10Higher prevalence rate of H. pylori in the elderly has been reported. 23Results of the current study showed a strong correlation between seropositivity rates and age increase (P < 0.05).Specific environmental factors linked to the higher age groups compared to aging are the major causes of this tendency. 23The present study also showed high prevalence of H. pylori seropositivity rates in younger age groups (52.7% at ≤30 years old and 40.9% at ≤20 years old) in Iran.The results showed anti-CagA seropositivity rates of 52.9% and 61.9% in seropositive and seronegative patients for anti-H.pylori antibody, respectively (P = 0.05).The CagA antibodies can be persevered in serum for a long period of time, implying the importance of these in diagnosis of H. pylori infections. 24These antibodies are important in diagnosing H. pylori infected patients who have sole positivity for anti-CagA antibodies, after seroconversion of general anti-H.pylori IgG.Simultaneous use of the serology test with an antigenbased stool test was recommended to detect the active H. pylori infections.

Conclusion
To the best of our knowledge, this is the first report on the high anti-CagA antibody in both seropositive and seronegative patients for general IgG, indicating the importance of this antibody in diagnosis of H. pylori infected patients after seroconversion of general anti-H.pylori IgG.The high prevalence of anti-H.pylori antibody, which was observed in dyspeptic population in this study, should also be considered in evaluating upper gastrointestinal diseases in Iran.

Limitations
We acknowledge that this study has some limitations.The temporality ascertainment of the present study and the number of patients investigated for anti-CagA antibody are the main limitations of this investigation, limiting its accuracy to show the true prevalence in country.More studies are needed on epidemiological and environmental factors.

Figure 1 .
Figure 1.The Rate of General Helicobacter pylori IgG and Anti-CagA IgG in Different Age Groups

Table 1 .
The Seroprevalence of Helicobacter pylori IgG Antibody in Iranian Dyspeptic Patients