Background: The rate of antibiotic resistance in Helicobacter pylori (H.pylori) is increasing in Vietnam in recent years and has become a main challenge in the management of H. pylori infection. Data about the resistance remains different depending on the time, demography and especially the methods of determining the antibiotics susceptibility. Aims: To determine the antibiogram of H.pylori strains isolated from a population in Central Vietnam to clarithromycin, levofloxacin by E-test and disk diffusion, compare two diffusion methods. Methods: 56 H.pylori strains were isolated from gastric biopsies of H.pylori infected patients from 7/2012 to 8/2013, of which 13 strains originated from patients in whom eradication of the infection failed after treatment. E-test was used to determine the minimum inhibitory concentrations (MICs) of clarithromycin (CH) and levofloxacin (LE). Disk diffusion was evaluated as an alternative method to determine susceptibility and compared with the E-test results. Results: In total, the resistant strains (regardless of previous eradication history) to CH, LE were 42.9% and 44.6%, respectively. The ratio of strains with secondary resistance was significantly greater than that of the strains with primary resistance, CH: 84.6% vs. 30.2%, LE: 61.5% vs. 39.5% (p < 0.05). The resistance rate to LE in female was significantly higher than in male (p < 0.05). All CH-sensitive strains by E-test had the inhibition diameters of CH was ≥ 24mm and all CH-resistant strains had the inhibition diameters was ≤ 18mm (breakpoint for MIC: 1µg/ml). To LE, the inhibition diameters was ≥ 30mm can determine all LE-sensitive strains and the inhibition diameters was ≤ 26mm can determine all LEresistant strains by E-test (breakpoint for MIC: 1µg/ml). Conclusions: High resistance rate to CH and LE, suggests that standard CH-based triple therapies may not be useful as the first-line treatment and LE-based triple therapy should not use as an alternative therapy in Central Vietnam. The disk diffusion can be used as an alternative phenotypic method to determine the susceptibility of H.pylori, which may be more feasible and less expensive.