Although H. pylori infections, association with disorders of the bowel, as well as the possible involvement of cancers in the gut is an issue that has been studied extensively in developed countries; nevertheless, it is detection and diagnosis in third world countries continue to be in accurate and controversial. This is probably related to the economic conditions and the poor health system in these countries. In Sudan and probably many of the third world countries the cost of diagnosis plays a major role, rather than the accuracy of the diagnostic method. Hence, diagnosis of H. pylori infections is largely based on serology, detection of stool antigen and rarely endoscopy and culture.
Currently there are many diagnostic methods for the diagnosis of H. pylori infections; each method has its advantages and disadvantages, so it is recommended to use at least a combination of two methods based on different principles to detect colonization by H. pylori (Ramis et al., 2012). Although, the culture method is regarded as the most appropriate technique; it has limitations particularly in case of slow-growing or fastidious bacteria, due to complicated identification and time-consuming methods. In addition to need for immediate transport of the biopsy specimens to the designated laboratory to assure the viability of H. pylori and prevent the formation of coccoid forms of the microorganism (Dong et al., 2019, Kisa et al., 2002, Ramis et al., 2012). The histological technique and culturing of gastric biopsy specimens have been considered a gold standard method under optimal conditions(Ramis et al., 2012).
Histological staining enables identifying bacteria and evaluating the type and intensity of the gastric mucosa's inflammation and associated pathology, such as, atrophic gastritis (AG), intestinal metaplasia (IM), and gastric cancer or lymphoma (Lee and Kim, 2015).
In this study, the prevalence of H. pylori infection was 35.5%. H. pylori was detected in (103/290) patients using histopathological examination with 35.5% sensitivity. There are many previous studies done in this field with various pictures of the disease. Mohamed et al. reported that 16/69 (23.2%) positive patients for H. pylori infection among Sudanese patients with colon polyps and colon cancer patients(Mohamed et al., 2020). Redéen et al. reported that 97/304 (31.9%) positive patients for H. pylori infection (Redéen et al., 2011). In another study Salman et al.,reported that 115/210 (54.7%) samples were positive for H. pylorivia histopathology, 57 (62.6%) of positive H. pylori samples observed in patients with chronic gastritis, 11 (50%) with adenocarcinoma and 31 (44.2%) with superficial gastritis, while only one H. pylori-positive out of 5 cases observed in atrophy gastritis patient (Salman et al., 2019).Histopathology is the first diagnostic method for detection of H. pylori and is still widely used as the main diagnostic tool; nevertheless, it has limitations including higher cost, longer turnaround time, and inter-observer variation assessment; experience and skills of the pathologist do matter for the specificity and sensitivity of histopathological diagnosis of H. pylori (Khalifehgholi et al., 2013), false positive results can occur due to presence of structures similar to H. pylori (Ramis et al., 2012)and failure todetect all the positive samples might occur in case of intestinal metaplasia (Salman et al., 2019). The density and irregular distribution of H. pylori can vary at different sites on the gastric mucosa which might lead to sampling error (Lee and Kim, 2015;Ramis et al., 2012). Moreover, the sensitivity of histology may decrease in patients taking antisecretory therapy, such as, proton pump inhibitor (PPI) (Lee and Kim, 2015).
Molecular tests should be applied as replacements to traditional method for the identification of H. pylori, which are a sensitive, rapid, and precise techniques for the specific recognition of H. pylori from gastric biopsy specimens and to discover particular mutations related to antimicrobial resistance (Dong et al., 2019;Kisa et al., 2002;Ramis et al., 2012).
In this study, identification of H. pylori was applied to all biopsies by PCR using specific primers. Specific H. pylori 16S rRNA gene is a conserved region of prokaryotic DNA that allows specific identification. However, H. pylori 16S rRNA gene's sensitivity and specificity were 46.6% and 78.6%, respectively. H. pylori ureA gene is an important virulence factor that ensures that bacteria can resist acidity of the gastric mucosa. ureA gene shows 35% sensitivity and 89.3% specificity. In our study, the glmM gene showed the lowest sensitivity (24.3%), and good specificity (92.5%). Our result aligned with a study conducted by AlNaji et al. 2018, which found that the glmM gene is 38.8% lower than the 16srRNA gene 95.9% (AlNaji et al., 2018). Helaly et al. reported similar results (38.5%) for glmM gene (Helaly GF et al., 2009). This low percent of glmM (ureC) gene may be due to sequence polymorphism or/in variation to the diversity of strains within the patients that reported in previous studies (AlNaji et al., 2018), also housekeeping genes are affected by geographical regions and point mutations, Intragenic and recombination are another potential factors (Raymond et al., 2004). The ureC gene encodes phosphoglucosamine mutase, which is unconnected to urease production and it is renamed by De Reuse et al. in 1997 to glmM (De Reuse et al., 1997).
TheureA gene is a housekeeping gene that is needed for urease enzyme activity. Espinoza et al demonstrated that the amplification of the ureA gene was noticed in (86.36%) which was lower than that of the glmM gene (100%) (Espinoza et al., 2011). Smith et al. reported that ureA gene PCR had a very poor specificity and sensitivity (Smith et al., 2004). The possible reasons for poor sensitivity of ureA and ureC (glmM) genes for detection of H. pylori may be that both of them are single-step PCR and thus unable to identify the lower number of bacteria or they were unable to counteract PCR inhibitors in the clinical specimens (Singh et al., 2008).
The 16S rRNA gene is a useful and commonly used for primary finding of H. pylori use Hp1, Hp2 primers with sensitivity up to 100% (AlNaji et al., 2018). Sugimoto et al. and Farhadkhani et al. reported that for the detection of H. pylori 16S rRNA gene was greater than ureA gene and they determined that the difference could be due to discrepancy in the primer specificity and sensitivity. Using of 16S rRNA gene for detection of H. pylori might be more sensitive, but could not be as specific as ureA gene (Farhadkhani et al., 2019;Sugimoto et al., 2009). The poor specificity may be explained by sequence conservation across the bacterial genera and also by possible amplification of nonspecifically human DNA (Singh et al., 2008). Yet, no 100% specificity or sensitivity for primer sets amplifies H. pylori ureA and 16S rRNA genes (Farhadkhani et al., 2019;Sugimoto et al., 2009).