摘要
目的:建立PCR直接检测临床血标本中病原菌的方法,探索其快速诊断全身性感染(sepsis)的应用价值.方法:利用16S rRNA基因的高度保守性,设计并合成细菌的通用引物;采用合成的引物扩增标准菌株及全身性感染病人的血样本,并与血培养进行对照.结果:通用引物PCR可检出0.005pg的大肠埃希菌标准菌株DNA;临床分离的大肠埃希菌、金黄色葡萄球菌PCR扩增产物于255bp处均可见清晰电泳条带,正常人无菌血白细胞及白色念珠菌DNA扩增产物均未见电泳条带出现;PCR可快速地检出血液中多种病原菌DNA,所检的8例病人中4例阳性(50%).结论:165 rRNA基因为基础的PCR,可直接用于临床血标本病原菌的检测,初步实验显示其具有特异、快速等优点,敏感性较血培养有增高趋势,有利于重症急性胰腺炎及其他外科危重病Sepsis的早期诊断.
Objective: To establish a PCR method which could be used to directly detect pathogenic bacterial components in the blood of patients with sepsis and to explore the value of this method diagnosis rapidly. Methods: A pair of universal primer for eubacteria targeting conserved region in 16S rRNA gene were designed. The primers were used to detect the blood sample of patients with sepsis. And the sensitivity and specificity of this method were tested. Results: DNA genes of Gram-positive Staphylococcus aureus and Gram-negative Escheria coli gave a visible band after amplification but fungi and white cell in healthy volunteers were negative. The sensitivity test showed that the PCR amplification technique could detect 0.005pg of genomic Escheria coli DNA. 50 percent of patients with sepsis had microbial DNA which was detected in their blood, but only one had positive blood cultures. Conclusions: PCR based on 16S rRNA may be directly used to detect pathogenic bacterial components in blood. The preliminary study showed that it is beneficial for early diagnosis of sepsis in SAP and other surgical critical ill patients. The sensitivity of PCR method showed a tendency towards more sensitive than blood cultures.
出处
《外科理论与实践》
1998年第2期84-87,共4页
Journal of Surgery Concepts & Practice
