BACKGROUND: The eradication rate of Helicobacter pylori is steadily decreasing because of increasing resistance to clarithromycin. According to the new version of Maastricht IV guidelines, molecular tests can be performed as a substitute for bacterial culture and the standard clarithromycin susceptibility test for the detection of H. pylori and clarithromycin resistance directly on gastric biopsy samples. OBJECTIVE: To evaluate the clinical efficacy of H. pylori detection using a molecular test and treatment outcomes of the clarithromycin-based genotypic resistance test. MATERIALS AND METHODS: A total of 385 patients diagnosed with functional dyspepsia were recruited in this clinical trial. Total DNA was extracted from formalin-fixed paraffin-embedded samples and prepared for a molecular test and H. pylori detection was performed simultaneously by modified Giemsa staining. Genotypically sensitive patients with positive H. pylori were treated by quadruple therapy: bismuth potassium citrate, rabeprazole, amoxicillin, and clarithromycin (BRAC) and genotypically resistant individuals were treated by bismuth potassium citrate, rabeprazole, amoxicillin, and furazolidone (BRAF) twice daily for 7 consecutive days. The eradication rate of H. pylori was assessed using the C-urea breath test at 6 weeks after treatment. RESULTS: The prevalence of H. pylori infection in functional dyspepsia patients was 35.3% (136/385), 29.1% for women (53/182) and 40.9% for men (83/203). The sensitivities of real-time PCR and histological examinations were 95.6% (130/136) and 69.9% (95/136). Forty-one samples were found to be positive by real-time PCR alone and six by histological examination alone, the majority of which (32/41, 5/6) were identified as grade 1 multiplicity of infection. The overall resistance rate to clarithromycin was 37.7% (49/130): 37.3% (19/51) for women and 38.0% for men (30/79). Eighty-nine patients with positive H. pylori detected by both real-time PCR and histological examinations received quadruple therapies. For the intention-to-treat analysis, the eradication rates of BRAC and BRAF were 98% (52/53) and 92% (33/36), or 100% (52/52) and 94% (33/35) for per-protocol analysis. CONCLUSION: Real-time PCR is efficacious for H. pylori detection and genotypic resistance-guided quadruple therapy has a high efficacy in treating functional dyspepsia with H. pylori infection.