Background: Helicobacter pylori infects approximately half of the global population and is the major cause of peptic ulcer disease. Chronic H. pylori infection is also a key risk factor for gastric adenocarcinoma and mucosa-associated lymphoid tissue lymphoma, which together account for ~850,000 new cancer cases worldwide each year (GLOBOCAN 2022; IARC). Increasing antibiotic resistance—particularly to clarithromycin and levofloxacin—compromises empiric eradication therapies and drives the need for comprehensive molecular susceptibility testing. Currently, routinely available assays are mostly qPCR-based and limited to a small number of resistance markers. Targeted next-generation sequencing (NGS) offers broader resistance profiling but remains largely inaccessible for routine use. Deeplex® HelP was developed as a ready-to-use targeted NGS assay to support susceptibility-guided management of H. pylori infection through multidrug resistance prediction directly from gastric biopsies.
Objectives: This study evaluates the clinical performance of Deeplex® HelP on gastric biopsies for the prediction of resistance to antibiotics commonly used in H. pylori eradication regimens.
Methods: A total of 198 H. pylori-positive gastric biopsies collected at Poitiers University Hospital were analysed with Deeplex® HelP. Briefly, multiplex PCR was performed to amplify key genomic regions associated with H. pylori resistance to clarithromycin, levofloxacin, rifampicin and tetracycline (23S, gyrA, gyrB, rpoB and 16S), followed by NGS and automated data analysis. Deeplex® HelP results were compared with standard phenotypic antimicrobial susceptibility testing (AST) and a commercially available qPCR assay for clarithromycin resistance prediction based on the 23S gene (Allplex™ H. pylori & ClariR Assay).
Results: Deeplex® HelP generated complete results for all four antibiotics in 190 of 198 biopsies, with qPCR Ct values ranging from 15 to 34. Four samples yielded partial results, and four others yielded no results. The latter had high Ct values (32–36), indicating that assay performance is affected only at very low bacterial loads. For clarithromycin and levofloxacin, Deeplex® HelP correctly identified 93.5% and 79.3% of resistant strains and 98.5% and 98.8% of susceptible strains, with heteroresistance detected in 19 and 11 samples. Clarithromycin resistance prediction by Deeplex® HelP and the 23S-based qPCR assay was concordant in 95.4% of cases. For rifampicin and tetracycline, all susceptible strains were correctly predicted. The two rifampicin-resistant strains were predicted susceptible, with no known rpoB variants detected, and AST is being repeated to confirm resistance. Overall, the low prevalence of resistant strains in this dataset is consistent with previously reported rates in the literature. Discordant cases are under investigation and may reflect limitations of culture-based AST, including missed heteroresistance or variability near EUCAST MIC breakpoints.
Conclusion: Deeplex Help® provides rapid, reliable detection of H. pylori resistance variants, including rare and minority ones, with high concordance for clarithromycin and levofloxacin. By overcoming limitations of conventional PCR and culture, it enables comprehensive resistance profiling to guide personalized eradication therapy.