焦磷酸测序法在败血症常见病原体检测中的应用.pdfVIP

J South Med Univ, 2013, 33(7): 1053-1056 doi 10.3969/j.issn. 1673-4254.2013.07.24 ·1053 · 基础研究 焦磷酸测序法在败血症常见病原体检测中的应用焦磷酸测序法在败血症常见病原体检测中的应用 1 2 1 1 胡子有 ，韩 慧 ，曾 勇 ，吴炳义 1 2 南方医科大学南方医院临床医学实验研究中心，广东 广州 510515 ；中山大学附属中山医院检验医学中心， 广东 中山 528403 摘要：目的将焦磷酸测序技术应用于败血症常见病原体的检测鉴定。方法 根据细菌16S rRNA序列的特点，设计焦磷酸测序 用的PCR 扩增引物和测序引物，提取败血症常见病原体的基因组DNA ，进行焦磷酸测序，分析获得的序列，进行比对，对病原体 种类作出判断。结果 经焦磷酸测序可以获取有效长度28 bp 的序列，使用该测序序列可以有效区分化脓性链球菌、肺炎链球 菌、大肠埃希菌、铜绿假单胞菌、肺炎克雷白杆菌、脑膜炎双球菌、沙门氏菌，但不能区分金黄色葡萄球菌和表皮葡萄球菌。结论 应用焦磷酸测序可以较有效地对败血症常见病原体的种类实现种属水平的区分。 关键词：焦磷酸测序；败血症；病原体；16sRNA Application of pyrosequencing in detection of common pathogens in sepsis 1 2 1 1 HU Ziyou , HAN Hui , ZENG Yong , WU Bingyi 1Research Center of Clinical Medicine, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China; 2Laboratory Medical Center, Zhongshan Hospital of Sun Yat-sen University, Zhongshan 528403, China Abstract: Objective To apply pyrosequencing technique in the detection of the common pathogens in sepsis. Methods The primers for amplification and sequencing in pyrosequencing were designed according to alignment of the bacterial 16S rRNA sequence. Bacterial genomic DNA was extracted for pyrosequencing, and the pathogen species were determined according to the sequencing data obtained. Results Pyrosequencing effectively yielded the sequencing data of the 28 bp sequences of the pathogens and clearly distinguished the pathogen species of Streptococcus pyogenes, Streptococcus pneumonia, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumonia, Neisseria meningitides, and Salmonella, but failed to distinguish Staphylococcus epidermidis from Staphylococcus aureus. Conclusion Pyrosequencing technique can effectively distinguish the common pathogens in se