实时荧光定量PCR技术在侵袭性真菌感染诊断中的价值

发布时间：2018-06-11 15:49

本文选题：侵袭性真菌感染 + RTFQ-PCR　；参考：《宁夏医科大学》2016年硕士论文

【摘要】：目的:本研究将实时荧光定量PCR技术(Real-time Flurescent Quantitive Polymerase Chain Reaction,RTFQ-PCR)应用于侵袭性真菌感染(Invasive fungal infection IFI)的早期诊断,旨在评价RTFQ-PCR技术在IFI早期诊断中的应用价值,为早期诊断与治疗IFI提供依据。方法:1、采集宁夏医科大学总医院及心脑血管病医院ICU科2015年5月至2016年3月IFI高危患者102份血清,同时收集其相关临床资料。2、送检血培养、G试验。3、RTFQ-PCR检测:(1)提取白色念珠菌DNA,经PCR扩增出目的基因片段并回收,然后倍比稀释,分别计算出拷贝数,最后经RTFQ-PCR检测,由软件绘制出标准曲线。(2)提取IFI高危患者血清中的DNA,进行RTFQ-PCR检测,收集实验数据。4、根据“重症患者IFI的诊断和治疗指南(2007年)”制定的诊断标准,将纳入的研究对象分为:确诊IFI、临床诊断IFI、拟诊IFI及非IFI。将“2007年指南”作为金标准,以确诊和临床诊断病例作为感染组,非IFI病例作为非感染组,将感染组和非感染组分别作为阳性对照病例和阴性对照病例,计算RTFQ-PCR和G试验的敏感度(Sensitivity Se)、特异度(Specificity Sp)、阳性预测值(Positive predictive value PPV)、阴性预测值(Negative predictive value NPV),分析评价RTFQ-PCR在IFI早期诊断中的应用价值。5、以血培养为金标准,将血培养阳性作为阳性对照病例,血培养阴性做为阴性对照病例,计算RTFQ-PCR的敏感度、特异度、阳性预测值、阴性预测值。结果:1、本次研究收集了102份标本,最终共纳入99份标本做为研究对象,其中,确诊IFI病例2例,临床诊断IFI病例20例,拟诊IFI病例28例,非IFI病例49例。以“2007年指南”作为金标准,计算RTFQ-PCR的敏感度为50%,特异度为96%,阳性预测值为82%,阴性预测值为84%;G试验的敏感度为54%,特异性为92%,阳性预测值为76%,阴性预测值为80%;联合应用RTFQ-PCR和G试验的敏感度为78%,特异度为88%,阳性预测值为70%,阴性预测值为91%。2、以血培养为金标准,计算RTFQ-PCR的敏感度为100%,特异度为88%,阳性预测值为15%,阴性预测值为100%。结论:1、真菌RTFQ-PCR检测技术可以作为重症患者IFI早期诊断的一种有效手段,在临床上对IFI的早期诊断中具有一定的价值。2、RTFQ-PCR和G实验的联合应用,综合诊断价值较高,可以改善IFI的早期诊断率。
[Abstract]:Objective: in this study, Real-time Flurescent quantitative Polymerase chain reaction (RTFQ-PCR) was applied to the early diagnosis of invasive fungal infection IFI, in order to evaluate the value of RTFQ-PCR in the early diagnosis of IFI and provide evidence for early diagnosis and treatment of IFI. Methods: from May 2015 to March 2016, 102 IFI high risk patients were collected from ICU department of Ningxia Medical University General Hospital and Cardiovascular and Cerebrovascular Disease Hospital. At the same time, the relevant clinical data. 2. Blood culture test. RTFQ-PCR was used to extract the DNA of Candida albicans. The target gene fragment was amplified by PCR and recovered, then the copy number was calculated by double dilution, and finally detected by RTFQ-PCR. Drawing out the standard curve by software, extracting the DNA from the serum of IFI high risk patients, detecting it by RTFQ-PCR, collecting the experimental data .4. according to the diagnostic criteria of "guidelines for the diagnosis and treatment of IFI in severe patients (2007)". The subjects were divided into four groups: confirmed IFI, clinical IFI, IFI and non IFI. The 2007 guidelines were used as the gold standard, with confirmed and clinically diagnosed cases as the infected group, non-IFI cases as the non-infected group, and the infected and non-infected groups as the positive control cases and the negative control cases, respectively. The sensitivity, specificity, positive predictive value and negative predictive value of RTFQ-PCR and G test were calculated to evaluate the sensitivity, specificity, positive predictive value and negative predictive value of RT FQ-PCR and G test. The value of RTFQ-PCR in the early diagnosis of IFI was evaluated. The sensitivity, specificity, positive predictive value and negative predictive value of RTFQ-PCR were calculated. Results in this study, 102 specimens were collected and 99 specimens were included in the study. Among them, 2 cases were confirmed IFI cases, 20 cases were clinically diagnosed IFI cases, 28 cases were IFI cases, 49 cases were non-IFI cases. Using the 2007 Guide as the gold standard, The sensitivity of RTFQ-PCR is 50, the specificity is 96, the positive predictive value is 82, the negative predictive value is 84g, the sensitivity is 54, the specificity is 92, the positive predictive value is 76and the negative predictive value is 80. The sensitivity of the combination of RTFQ-PCR and G test is 78. The specificity is 88. The positive predictive value is 70. The negative predictive value is 91. 2. Blood culture is the gold standard. The sensitivity, specificity, positive predictive value and negative predictive value of RTFQ-PCR were 100, 88, 15 and 100, respectively. Conclusion the detection of fungal RTFQ-PCR can be used as an effective method for the early diagnosis of IFI in severe patients. It has a certain value in the early diagnosis of IFI. The combined use of RTFQ-PCR and G test is of great value in the clinical diagnosis of IFI, and the comprehensive diagnostic value is higher. It can improve the early diagnosis rate of IFI.
【学位授予单位】：宁夏医科大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R440;R519

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