【摘要】：目的：侵襲性曲霉菌感染（invasive aspergillosis，IA）是一種嚴重威脅免疫力低下病人生命的感染，早期準確的診斷和治療是良好預后的關鍵。本研究首先建立核酸序列依賴擴增（nucleic acid sequence-basedamplification，，NASBA）檢測曲霉菌的方法，再結合熒光定量PCR（qPCR）及GM試驗應用于IA的臨床診斷，確立IA的最佳實驗室診斷策略。 方法：首先在實驗室內利用煙曲霉菌標準株（CMCC A1a）建立NASBA結合電泳檢測曲霉菌的方法，并對其靈敏度、特異性進行了評估。然后對NASBA結合分子信標（MB）用于定量檢測曲霉菌的可行性進行了探索。最后分別利用NASBA、熒光定量PCR及GM試驗三種方法對80例IA高危病患的血液標本進行檢測，計算三種診斷方法的各項性能指標，并對各種診斷方案進行比較，最終確定IA實驗室診斷的最佳方案。 結果：在實驗室內成功建立NASBA檢測曲霉菌的方法，其特異性良好，靈敏度達到了1Cfu；曲霉菌孢子量的對數(shù)與MB熒光信號達到設定閾值時的時間存在線性相關關系（y=-10.7x+81.6，r=0.9889）。 NASBA、qPCR及GM試驗三種方法的靈敏度分別為76.47%、67.65%、52.94%，特異度分別為80.43%、89.13%、80.43%。聯(lián)合診斷結果顯示，NASBA與qPCR串聯(lián)方案有最好的特異度（100%）及陽性預測值（100%）；NASBA與qPCR并聯(lián)方案則最為靈敏（94.12%）。 結論：NASBA結合MB可用于定量檢測曲霉菌。NASBA用于診斷IA最為敏感，而qPCR則最為特異，GM試驗的表現(xiàn)差于前兩者。聯(lián)合應用的診斷策略能夠提高診斷效能，提高臨床早期診斷IA的準確性。
[Abstract]:Objective: invasive aspergillus infection (invasive aspergillosis IA) is a serious threat to the life of patients with low immunity infection, early accurate diagnosis and treatment is the key to a good prognosis. In this study, a method for detecting Aspergillus by sequence-dependent amplification (nucleic acid sequence-amplified amplification (NASBA) was established, and then the best laboratory diagnostic strategy was established by using fluorescence quantitative PCR (qPCR) and GM test in the clinical diagnosis of IA. Methods: first of all, the standard strain of Aspergillus fumigatus (CMCC A1a) was used to establish a method for the detection of Aspergillus fumigatus by NASBA combined with electrophoretic electrophoresis, and its sensitivity and specificity were evaluated. Then the feasibility of quantitative detection of Aspergillus by NASBA combined with molecular beacon (MB) was explored. Finally, three methods, NASBA, fluorescent quantitative PCR and GM test, were used to detect the blood samples of 80 patients with high risk of IA, to calculate the performance indexes of the three diagnostic methods, and to compare the various diagnostic schemes. Finally, the best scheme for IA laboratory diagnosis was determined. Results: the method of detecting Aspergillus by NASBA was successfully established in laboratory. The specificity of the method was good and the sensitivity reached 1Cfu. There was a linear correlation between the logarithm of aspergillus spores and the time when the MB fluorescence signal reached the threshold (Y-10.7x 81.6rr = 0.9889). The sensitivity of NASBA qPCR and GM test was 76.477.65 and 52.94, respectively, and the specificity was 80.4389.133.The specificity of the three methods was 80.43 89. 13 and 80.43, respectively. The results of combined diagnosis showed that the best specificity (100%) and positive predictive value (100%) of NASBA and qPCR tandem protocol were the most sensitive (94.12%). Conclusion the ratio NASBA combined with MB can be used to detect aspergillus. NASBA is the most sensitive in the diagnosis of IA, while qPCR is the most specific for GM test than the former two. The combined diagnostic strategy can improve the diagnostic efficacy and improve the accuracy of early diagnosis of IA.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R519;R446

【參考文獻】

相關期刊論文 前2條

1 孟浦;肖晶;潘峰;聶秀;周東風;何世斌;李立家;;FISH檢測快速診斷曲霉菌感染的臨床價值[J];中國組織化學與細胞化學雜志;2010年04期

2 黎庶;張立娜;王欣;劉婷;李佩玲;謝秀麗;徐克;;侵襲性肺曲霉菌感染CT征象的早期表現(xiàn)及其隨訪觀察[J];中國臨床醫(yī)學影像雜志;2009年08期

本文編號：2151480