Novel chromogenic medium-based method for the rapid detection of Helicobacter pylori drug resistance

Helicobacter pylori (H. pylori), a globally prevalent pathogen, is exhibiting increasing rates of antimicrobial resistance. However, clinical implementation of pre-treatment susceptibility testing remains limited due to the organism's fastidious growth requirements and prolonged culture time.

To propose a novel detection method utilizing antibiotic-supplemented media to inhibit susceptible strains, while resistant isolates were identified through urease-mediated hydrolysis of urea, inducing a phenol red color change for visual confirmation.

Colombia agar was supplemented with urea, phenol red, and nickel chloride, and the final pH was adjusted to 7.35. Antibiotic-selective media were prepared by incorporating amoxicillin (0.5 μg/mL), clarithromycin (2 μg/mL), metronidazole (8 μg/mL), or levofloxacin (2 μg/mL) into separate batches. Gastric antral biopsies were homogenized and inoculated at 1.0 × 10⁵ CFU onto the media, and then incubated under microaerobic conditions at 37 °C for 28-36 hours. Resistance was determined based on a color change from yellow to pink, and the results were validated via broth microdilution according to Clinical and Laboratory Standards Institute guidelines.

After 28-36 hours of incubation, the drug-resistant H. pylori isolates induced a light red color change in the medium. Conversely, susceptible strains (H. pylori 26695 and G27) produced no visible color change. Compared with the conventional 11-day protocol, the novel method significantly reduced detection time. Among 201 clinical isolates, 182 were successfully evaluated using the new method, resulting in a 90.5% detection rate. This was consistent with the 95.5% agreement rate observed when compared with microdilution-based susceptibility testing. The success rate of the novel approach was significantly higher than that of the comparative method (P < 0.01). The accuracy of the new method was comparable to that of the dilution method.

The novel detection method can rapidly detect H. pylori drug resistance within 28-36 hours. With its operational simplicity and high diagnostic performance, it holds strong potential for clinical application in the management of H. pylori antimicrobial resistance.