Related Experiment Video
Updated: Jun 19, 2025
Gastric Mucosa Quantitative Polymerase Chain Reaction Analysis for Detecting Helicobacter pylori and Antibiotic Resistance
Published on: March 7, 2025
Simultaneous Detection of Helicobacter pylori and Clarithromycin Resistance Mutations Using RAA-CRISPR/Cas13a Assay
Kuocheng Yan1, Xiaohui Wang2, Yao Han1
1State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, People's Republic of China.
Background:
Infection caused by Helicobacter pylori (H. pylori) affects approximately 50% of the global population. It is a major pathogenic factor for chronic gastritis and gastric cancer. Besides, the resistance to antibiotics such as clarithromycin could reduce the eradication rate. Currently, there is an urgent need for a swift, easy to perform, and highly sensitive detection method for H. pylori and clarithromycin resistance.
Methods:
We used FAM/Digoxin labeled primers to amplify specific H. pylori 23S rRNA fragments by Recombinase Aided Amplification (RAA), and resistance mutations were distinguished using CRISPR/Cas13a system combined with lateral flow strip. Twenty-eight saliva samples were analyzed using qPCR, gene sequencing and this method to evaluate the detection efficiency.
Results:
We developed a simultaneous detection method for H. pylori and clarithromycin resistance mutations named sensitive H. pylori easy-read dual detection (SHIELD). The results showed both A2142G and A2143G mutant DNAs causing clarithromycin resistance could be distinguished from the wild type with a concentration of 50 copies/μL, and no cross-reaction with other 5 common gastrointestinal bacteria was observed. For the detection of H. pylori in 28 saliva samples, the positive predictive value of this method was 100% (19/19) in comparison with qPCR. For detecting clarithromycin resistance, the positive predictive value of this method was 84.6% (11/13) compared with gene sequencing.
Conclusion:
SHIELD assay showed high sensitivity and specificity in detecting H. pylori and clarithromycin resistance mutations. It could be a potential measure in the rapid detection of H. pylori, large-scale screening and guiding clinical medication.
