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目的探讨儿童化脓性脑膜炎(化脑)新的快速诊断方法。方法2003-08—2005-12采用16SrRNA荧光定量法对浙江大学儿童医院49例临床疑似化脑患儿脑脊液(CSF)的细菌DNA进行测定;监测化脑患儿脑脊液细菌DNA拷贝数,同期进行CSF细菌培养的对照。结果(1)荧光定量PCR(FQ-PCR)检测49份脑脊液标本发现17份阳性,阳性率为34.7%(17/49),明显高于脑脊液培养的阳性率10.2%(5/49),差异具有显著性(P<0.01)。(2)对17份FQ-PCR阳性标本进一步测定细菌DNA的拷贝数,发现患儿病情与其DNA拷贝数呈正相关,与其Ct值(指每个反应管内的荧光信号到达设定的阈值时所经历的循环数)呈负相关,Ct值越低,脑脊液细菌DNA拷贝数越高,患儿的预后越差。(3)FQ-PCR、CSF细菌培养同时阳性的仅为5例。(4)对2例脑脊液FQ-PCR的产物测序,Ct值17.9的测序提示为大肠埃希菌,符合CSF细菌培养结果;Ct值31.8的,测序未果。结论荧光定量PCR特异性强、敏感性高,需标本量少,是早期快速诊断儿童化脑的可靠方法,具有较大的应用价值。
Objective To explore a new rapid diagnostic method for purulent meningitis in children. Methods 2003-08-2005-12 Bacterial DNA of cerebrospinal fluid (CSF) from 49 cases of clinical suspected cerebral peduncle in Children’s Hospital of Zhejiang University was measured by 16S rRNA fluorescence quantitative method. The number of bacterial DNA in cerebrospinal fluid Bacterial culture control. Results (1) FQ-PCR detected 17 positive samples in 49 cerebrospinal fluid samples, the positive rate was 34.7% (17/49), significantly higher than that of cerebrospinal fluid culture (10.2%, 5/49) Significant (P <0.01). (2) To further determine the copy number of bacterial DNA in 17 FQ-PCR positive specimens, we found that the condition of children was positively correlated with their DNA copy number. Compared with the Ct value (the fluorescence signal in each reaction tube reached the set threshold The number of cycles) was negatively correlated, the lower the Ct value, the higher the bacterial DNA copy number in CSF was and the poorer the prognosis in children was. (3) FQ-PCR, CSF positive bacterial culture only 5 cases. (4) FQ-PCR products were sequenced in 2 cases of cerebrospinal fluid. The Ct value of 17.9 was identified as Escherichia coli, which accorded with CSF bacterial culture results. The Ct value of 31.8 was not detected. Conclusion Fluorescent quantitative PCR has the characteristics of high specificity, high sensitivity and small amount of samples, which is a reliable method for early diagnosis of child brain. It is of great value in application.