Columbia blood plate

According to the ‘National Clinical Laboratory Procedures’, routine testing methods were utilized. Drug susceptibility test was performed by paper diffusion method (K-B method). CLSI 2008–2010 operation procedures and result evaluation standards were used. Columbia blood plate, MH plates, identifying culture medium and drug sensitive slips were provided by the Oxoid (London, UK). The quality control strains viz. Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27853 and Escherichia coli ATCC 25922 were provided by provincial CDC clinical laboratory test center. The KB method is mainly used for the detection of ampicillin, cefotaxime, cefepime, ceftazidime, imipenem, clindamycin, vancomycin, erythromycin, roxithromycin, gentamicin and amikacin. The corresponding biological reagents were procured from Hangzhou Tianhe Microoragnism Reagents Co., Ltd. (Hangzhou, China). MH agar plates were stored at 4°C before use. Technicians with >5 years laboratory experience had performed all operations in strict accordance with the instructions.

Thawed the gastric mucosal specimen at room temperature and fully ground it into homogenate with 600 μL of brain heart infusion in a glass grinder. Spread the tissue homogenate on a Columbia blood plate (OXOID, England) containing 5% sheep blood by streaking. Cultivated the H. pylori in a 37 °C mixed gas incubator (5% O₂, 10% CO₂ and 85% N₂) for 2-3 d. Extended the culture time to 7 d if there was no growth. Picked out and subcultured the monoclonal colonies with typical colony morphology. Then smeared microscopy with the suspected H. pylori strain and carried out the biochemical experiments with urease (Haibo Biotechnology Co., Ltd., China), oxidase (Beijing Luqiao Technology Co., Ltd., China) and catalase (Haibo Biotechnology Co., Ltd., China). Suspected strains with biological morphology and biochemical reactions were identified as H. pylori, and the patient was diagnosed as positive for H. pylori infection. Extended the culture time to 7 d, and if there was still no H. pylori strain growth on the medium, the patient was diagnosed as negative for H. pylori infection.