葡萄球菌分类与鉴定的两种方法研究与探讨

目的 鉴于葡萄球菌传统表型鉴定方法的局限性,为确保药品微生物检验中葡萄球菌检验结果的准确性,依据《中国药典》2015年版四部通则,利用DNA测序技术对15株菌株成功进行了序列测定,并与传统的生化鉴定方法进行比较和分析。方法 用PCR仪扩增提取到的葡萄球菌基因组DNA,然后进行琼脂糖凝胶电泳并对目的条带进行切胶回收,用ABI 3500测序仪对15株菌株进行DNA序列测序,对测定结果用CExpress软件进行拼接,并与数据库进行比对,用该结果与VITEKⅡ生化鉴定仪的结果进行比较和分析。结果 15株菌株的测序结果除了10号菌株DNA相似度为93.38%外,其余均大于99%,序列比对结果与VITEKⅡ生化鉴定仪的结果一致。结论 利用DNA测序技术对葡萄球菌进行鉴定的结果比传统的表型鉴定和生化鉴定结果更为准确可靠。

Research on Two Methods of Classification and Identification of Staphylococcus

OBJECTIVE To sequence 15 strains of Staphylococcus by DNA sequencing technology according to the general principles of the Chinese Pharmacopoeia 2015 edition, and compare with traditional biochemical identification method in view of the limitations of traditional phenotypic identification method, in order to ensure the accuracy of the results of Staphylococcus detection in drug microbiological testing. METHODS The genomic DNA extracted from Staphylococcus was amplified by PCR, then agarose gel electrophoresis was performed and the target strip was recovered by gel cutting. DNA sequencing of 15 strains was carried out by ABI 3500 sequencer. The results were spliced by CExpress software and compared with the database. The results were compared and analyzed. RESULTS The similarity of the 15 strains shown by the sequencing results were more than 99% except strain 10 for which the DNA similarity was 93.38%. The results of sequence alignment were consistent with those of VITEK II biochemical analyzer. CONCLUSION DNA sequencing is more accurate and reliable than traditional phenotypic and biochemical identification for the identification of Staphylococcus.