【摘要】：目的:檢測近平滑念珠菌、熱帶念珠菌對FCZ、AMB的藥物敏感性，體外誘導近平滑念珠菌、熱帶念珠菌的FCZ耐藥株及AMB耐藥株，并檢測誘導的耐藥株中耐藥基因的表達情況，探討其對FCZ、AMB耐藥的分子機制。 方法：先以MTT微量液基稀釋法檢測近平滑念珠菌、熱帶念珠菌對FCZ、AMB的藥物敏感性，再用FCZ、AMB濃度遞增法體外誘導其耐藥株,并用實時熒光定量PCR法檢測野生株與耐藥株耐藥基因的表達水平。 結果：近平滑念珠菌、熱帶念珠菌的野生株有CYP51A、CYP51B、CDR1、CDR2、MDR1、MDR2、Yap1、Sho1mRNA低水平表達，，而近平滑念珠菌、熱帶念珠菌的AMB耐藥株中CYP51A、CYP51B、CDR1、CDR2、MDR1、MDR2、Yap1、Sho1基因的相對表達量明顯上升，差異有統(tǒng)計學意義（P＜0.05）；近平滑念珠菌、熱帶念珠菌的FCZ耐藥株中CYP51A、CDR1、CDR2、MDR1、Yap1、Sho1基因的相對表達量明顯高于其野生株，差異有統(tǒng)計學意義（P＜0.05），而CYP51B、MDR2基因的表達量在近平滑念珠菌、熱帶念珠菌FCZ耐藥株誘導形成過程中無明顯變化（P0.05）。 結論：CYP51A、CYP51B、CDR1、CDR2、MDR1、MDR2、Yap1、Sho1基因過度表達與近平滑念珠菌、熱帶念珠菌對AMB耐藥有關；CYP51A、MDR1、CDR1、CDR2、Yap1、Shol基因的過表達與近平滑念珠菌、熱帶念珠菌對FCZ耐藥有關，而近平滑念珠菌、熱帶念珠菌FCZ耐藥株誘導形成過程中CYP51B、MDR2基因的表達量無明顯變化。近平滑念珠菌、熱帶念珠菌長期接觸FCZ、AMB后可誘導麥角固醇合成過程中的關鍵酶基因、外排泵基因及信號傳導相關基因過表達，從而產生耐藥性。
[Abstract]:Objective: to detect the drug sensitivity of Candida albicans and Candida tropicalis to FCZ,AMB, to induce Candida meliformis, FCZ-resistant strains and AMB-resistant strains of Candida tropicalis in vitro, and to detect the expression of drug-resistant genes in the drug-resistant strains induced by Candida tropicalis and Candida tropicalis. To investigate the molecular mechanism of its resistance to FCZ,AMB. Methods: MTT microdilution method was used to detect the drug sensitivity of Candida solanum and Candida tropicalis to FCZ,AMB, and then the drug-resistant strains were induced by increasing the concentration of FCZ,AMB in vitro. Real-time fluorescence quantitative PCR was used to detect the expression level of drug-resistant genes in wild and drug-resistant strains. Results: low level expression of CYP51A,CYP51B,CDR1,CDR2,MDR1,MDR2,Yap1,Sho1mRNA was found in wild strains of Candida tropicalis and Candida tropicalis, while CYP51A,CYP51B,CDR1,CDR2,MDR1,MDR2,Yap1, was found in AMB resistant strains of Candida solani and Candida tropicalis. The relative expression of Sho1 gene increased significantly (P < 0.05). The relative expression of CYP51A,CDR1,CDR2,MDR1,Yap1,Sho1 gene in FCZ resistant strains of Candida leiomycetes and Candida tropicalis was significantly higher than that of wild strains (P < 0.05), while CYP51B, was significantly higher than that of wild strains (P < 0.05). The expression of MDR2 gene did not change significantly during the induction of drug-resistant strains of Candida moniliforme and Candida tropicalis FCZ (P0.05). Conclusion: the overexpression of CYP51A,CYP51B,CDR1,CDR2,MDR1,MDR2,Yap1,Sho1 gene is related to the drug resistance of Candida solanum and Candida tropicalis to AMB. The overexpression of CYP51A,MDR1,CDR1,CDR2,Yap1,Shol gene was related to the resistance to FCZ of Candida solanum and Candida tropicalis, but the expression of CYP51B,MDR2 gene did not change during the induction of drug resistance of Candida tropicalis and Candida tropicalis FCZ. Long-term exposure to FCZ,AMB can induce the over-expression of key enzyme genes, efflux pump genes and signal transduction-related genes in ergosterol synthesis in Candida leiomycetes and Candida tropicalis, resulting in drug resistance.
【學位授予單位】：福建醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R519.3

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