Determination of Helicobacter pylori cagA, vacA genotypes with real-time PCR melting curve analysis

Abstract

Objectives: genotypes of Helicobacter pylori are the focus of interest because they play a prominent role in mucosal injury. The purpose of this study was to determine cagA and vacA genotypes of H. pylori using real-time polymerase chain reaction (PCR) method with a double strain DNA binding SYBR Green I.dye, and to compare this with those of two immunohistochemical methods. Methods: forty-three paraffin-embedded biopsy tissue samples were examined by histology, epidermal growth factor receptor (EGFR), proliferating cell nuclear antigen (PCNA) immunohistochemistry and melting curve analysis of real-time PCR. Results: the presence of cagA gene was associated with a significantly higher frequency of gastritis ( P=0.003) than that of vacA gene with intestinal metaplasia ( P=0.045). Significant difference was found between the presence of cagA gene and EGFR expression in intestinal metaplasia cases in comparison with cagA negative samples ( P=0.0418). Statistically significant difference was detected between increased cell proliferation and the presence of gastritis. Conclusions: this method seems to be suitable for H. pylori genotype determination. Sensitivity, speed and simplicity are key areas in the development of PCR assays for H. pylori. Results supported the notion that infection with cagA positive H. pylori strain causes more augmentated cell proliferation in the stomach mucosa.