Gastric Mucosa Quantitative Polymerase Chain Reaction Analysis for Detecting Helicobacter pylori and Antibiotic Resistance.

Abstract

Helicobacter pylori is a main pathogen that infects nearly half of the global population and is threatening public health due to its increasing antibiotic resistance. Besides, Helicobacter pylori is also responsible for chronic gastritis, gastric and duodenal ulcers, gastric carcinoma, and gastric mucosa-associated lymphoid tissue (MALT) lymphoma. Therefore, it is essential to perform a timely and accurate diagnosis of H. pylori and the determination of its antibiotic resistance. Nowadays, existing methods of H. pylori diagnosis mainly include the rapid urease test (RUT), the urea breath test (UBT), the serum antibody test, the antigen test, gastroscopy, and bacterial culture. However, bacteria could not be cultured through the first five detection methods, not to mention the detection of drug resistance. The bacterial culture is time-consuming, and antibiotic sensitivity tests cannot be carried out rapidly and routinely. In clinical settings, the swift and precise identification of H. pylori and its susceptibility to antibiotics is crucial for its effective elimination. The objective of this protocol is to outline a targeted approach utilizing quantitative polymerase chain reaction (qPCR) on gastric mucosal samples to expedite the diagnosis of H. pylori and assess its resistance to antimicrobial agents. qPCR was exploited to detect the ureA gene for H. pylori infection and mutations in the 23S rRNA and gyrA genes associated with resistance to clarithromycin and quinolones, respectively. Currently, there remain challenges in gastric mucosa qPCR due to the lack of standard operating procedures. Therefore, it is essential to share methodologies with experimental details to ensure accurate communication of experimental procedures, contributing to gold-standard protocols that enable greater transparency. Overall, this protocol offers an economical and expeditious alternative to conventional methods for assessing H.pylori infection and its resistance to antibiotics through the application of quantitative polymerase chain reaction (qPCR) technology.