復發(fā)性外陰陰道白假絲酵母菌病的耐藥機制研究
發(fā)布時間：2018-11-06 07:32
【摘要】：目的：①對復發(fā)性外陰陰道假絲酵母菌病(Recurrent Vulvovaginal candidiasis,RWC)患者的致病白假絲酵母菌進行7種抗真菌藥物的體外藥物敏感實驗,明確RWC致病白假絲酵母菌藥物敏感性的差異；②對RVVC致病白假絲酵母菌進行藥物外排泵相關蛋白-多藥耐藥基因MDR1、三磷酸腺苷結合轉運蛋白家族CDR1、CDR2和PDR1基因表達產(chǎn)物的研究,明確上述耐藥基因在RVVC耐藥機制中的作用。 方法：①所有243株致病菌株均取自我院外陰陰道假絲酵母菌病患者,其中RVVC (實驗組)致病菌株84株,VVC(對照組)致病菌株159株,均以科瑪嘉顯色培養(yǎng)基、安圖顯色培養(yǎng)基及生物梅里埃VITEK2系統(tǒng)鑒定明確菌種；②以FUNGUS-7真菌藥敏試劑盒對已確定的213株白假絲酵母菌致病菌株進行2種多烯類藥物、4種唑類藥物和5-氟胞嘧啶共七種抗真菌藥的體外藥物敏感試驗；③以熒光定量PCR (realtime fluores-cence quantitative PCR, RTFQ PCR)對已確定的敏感、中敏和耐藥白假絲酵母菌菌株進行MDR1、CDR1、CDR2和PDR1基因表達產(chǎn)物的研究。 結果：①RVVC組白假絲酵母菌占90.48%,非白假絲酵母菌占9.52%,VVC組白假絲酵母菌占88.68%,非白假絲酵母菌占11.32%,兩組白假絲酵母菌構成比沒有顯著差異(P0.05)；②RWC組和VVC組致病白假絲酵母菌的體外藥敏結果顯示：RVVC組致病白假絲酵母菌對7種抗真菌藥物的敏感性依次為：制霉菌素兩性霉素B(5-氟胞嘧啶益康唑咪康唑氟康唑伊曲康唑；VVC組致病白假絲酵母菌對7種抗真菌藥物的敏感性依次為：兩性霉素B制霉菌素5-氟胞嘧啶咪康唑益康唑氟康唑伊曲康唑；IRVVC組對制霉菌素和咪康唑的敏感性顯著高于VVC組(P0.05)；③分別比較RVVC組和VVC組敏感、中敏、耐藥菌株中耐藥基因CDR1、CDR2、MDR1、PDR1的表達差異,結果顯示：PDR1基因不是在所有菌株均有表達,RWC組PDR1基因表達率明顯低于VVC組(P0.05)；PDR1基因的表達量比較顯示：5-氟胞嘧啶的WC-R組的表達量高于VVC-S組和VVC-I組(P0.05),兩性霉素B的VVC-R組高于VVC-I組(P0.05),伊曲康唑的RWC-R組高于WC-S組、WC-R組和RVVC-S組(P0.05);CDR1、CDR2、MDR1在所有白假絲酵母菌中均有表達,僅有CDR1基因表達量存在差異：5-氟胞嘧啶RVVC-R組明顯高于WC-R組(P0.05),咪康唑中敏、耐藥合并組VVC-I+R組高于VVC-S組(P0.05),氟康唑VVC-I組表達高于VVC-S組和VVC-R組(P0.05),但VVC-S組和WC-R組未檢出差異(P0.05)。 結論：①復發(fā)性外陰陰道假絲酵母菌病的主要致病菌仍為白假絲酵母菌；②顯色培養(yǎng)基適于臨床上白假絲酵母菌的快速鑒定,但對非白假絲酵母鑒定準確度有限;③RVVC和VVC致病白假絲酵母菌對抗真菌藥的敏感性大體一致,RVVC治療首選制霉菌素；④PDR1、CDR1基因可能為RWC和VVC患者致病白假絲酵母菌的耐藥相關基因；⑤除了耐藥基因表達增加以外,RWC耐藥機制的發(fā)生可能存在其它更重要相關因素。
[Abstract]:Objective: 1 to study the drug sensitivity of 7 antifungal drugs in patients with recurrent vulvovaginal Candida albicans (Recurrent Vulvovaginal candidiasis,RWC), and to determine the difference of drug sensitivity of RWC pathogenic Candida albicans. 2 the expression products of adenosine triphosphate transporter family (CDR1,CDR2) and PDR1 gene of drug efflux pump associated protein-multidrug resistance gene (MDR1,) were studied on Candida albicans caused by RVVC. To clarify the role of the above-mentioned drug resistance genes in the mechanism of drug resistance in RVVC. Methods: 1 all 243 strains of pathogenic bacteria were collected from patients with vulvovaginal candidiasis, including 159 strains of RVVC (experimental group, 84 strains of, VVC (control group). Antu chromogenic medium and biological Meridier VITEK2 system were used to identify the identified strains. (2) two polyenes, four azolides and seven 5-fluorocytosine antifungal agents were tested with FUNGUS-7 fungal susceptibility kit to 213 strains of Candida albicans. (3) the expression products of MDR1,CDR1,CDR2 and PDR1 genes were studied by fluorescence quantitative PCR (realtime fluores-cence quantitative PCR, RTFQ PCR) on the sensitive, moderately sensitive and drug-resistant Candida albicans strains. Results: in 1RVVC group, 90.48% of Candida albicans, 9.52% of non-Candida cerevisiae, 88.68% of Candida albicans, 11.32% of non-white Candida cerevisiae. There was no significant difference in the composition ratio of Candida albicans between the two groups (P0.05). In vitro antimicrobial susceptibility of pathogenic Candida albicans in 2RWC group and VVC group showed that the susceptibility of RVVC group to seven antifungal agents was nystatin amphotericin B (5-fluorocytosine econzomidazole). Conazole fluconazole itraconazole; The sensitivities of pathogenic Candida albicans to 7 antifungal agents in VVC group were amphotericin B nystatin 5 fluconazole miconazole econazole and itraconazole. The sensitivity of IRVVC group to nystatin and miconazole was significantly higher than that of VVC group (P0.05). (3) the difference of CDR1,CDR2,MDR1,PDR1 expression between RVVC group and VVC group was compared. The results showed that PDR1 gene was not expressed in all strains. The expression rate of PDR1 gene in RWC group was significantly lower than that in VVC group (P0.05). The expression of PDR1 gene in 5-fluorocytosine WC-R group was higher than that in VVC-S and VVC-I group (P0.05), and that in amphotericin B VVC-R group was higher than that in VVC-I group (P0.05). Itraconazole in RWC-R group was higher than that in WC-S group, WC-R group and RVVC-S group (P0.05). CDR1,CDR2,MDR1 was expressed in all Candida albicans, only CDR1 gene expression was different: 5-fluorocytosine RVVC-R group was significantly higher than WC-R group (P0.05), miconazole was sensitive to miconazole. VVC-I R group was higher than VVC-S group (P0.05), fluconazole VVC-I group was higher than VVC-S group and VVC-R group (P0.05), but there was no difference between VVC-S group and WC-R group (P0.05). Conclusion: 1 the main pathogenic bacteria of recurrent vulvovaginal candidiasis are still Candida albicans; (2) the chromogenic medium was suitable for rapid identification of Candida albicans in clinic, but the accuracy of identification for non-Candida albicans was limited, the sensitivity of antifungal agents against Candida albicans caused by 3RVVC and VVC was generally the same, and nystatin was the first choice in the treatment of RVVC. 4PDR1 + CDR1 gene may be the drug-resistance related gene of Candida albicans in RWC and VVC patients, and besides the increase of drug resistance gene expression, there may be other more important related factors in the mechanism of drug resistance of RWC.
【學位授予單位】：昆明醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R711.31
[Abstract]:Objective: 1 to study the drug sensitivity of 7 antifungal drugs in patients with recurrent vulvovaginal Candida albicans (Recurrent Vulvovaginal candidiasis,RWC), and to determine the difference of drug sensitivity of RWC pathogenic Candida albicans. 2 the expression products of adenosine triphosphate transporter family (CDR1,CDR2) and PDR1 gene of drug efflux pump associated protein-multidrug resistance gene (MDR1,) were studied on Candida albicans caused by RVVC. To clarify the role of the above-mentioned drug resistance genes in the mechanism of drug resistance in RVVC. Methods: 1 all 243 strains of pathogenic bacteria were collected from patients with vulvovaginal candidiasis, including 159 strains of RVVC (experimental group, 84 strains of, VVC (control group). Antu chromogenic medium and biological Meridier VITEK2 system were used to identify the identified strains. (2) two polyenes, four azolides and seven 5-fluorocytosine antifungal agents were tested with FUNGUS-7 fungal susceptibility kit to 213 strains of Candida albicans. (3) the expression products of MDR1,CDR1,CDR2 and PDR1 genes were studied by fluorescence quantitative PCR (realtime fluores-cence quantitative PCR, RTFQ PCR) on the sensitive, moderately sensitive and drug-resistant Candida albicans strains. Results: in 1RVVC group, 90.48% of Candida albicans, 9.52% of non-Candida cerevisiae, 88.68% of Candida albicans, 11.32% of non-white Candida cerevisiae. There was no significant difference in the composition ratio of Candida albicans between the two groups (P0.05). In vitro antimicrobial susceptibility of pathogenic Candida albicans in 2RWC group and VVC group showed that the susceptibility of RVVC group to seven antifungal agents was nystatin amphotericin B (5-fluorocytosine econzomidazole). Conazole fluconazole itraconazole; The sensitivities of pathogenic Candida albicans to 7 antifungal agents in VVC group were amphotericin B nystatin 5 fluconazole miconazole econazole and itraconazole. The sensitivity of IRVVC group to nystatin and miconazole was significantly higher than that of VVC group (P0.05). (3) the difference of CDR1,CDR2,MDR1,PDR1 expression between RVVC group and VVC group was compared. The results showed that PDR1 gene was not expressed in all strains. The expression rate of PDR1 gene in RWC group was significantly lower than that in VVC group (P0.05). The expression of PDR1 gene in 5-fluorocytosine WC-R group was higher than that in VVC-S and VVC-I group (P0.05), and that in amphotericin B VVC-R group was higher than that in VVC-I group (P0.05). Itraconazole in RWC-R group was higher than that in WC-S group, WC-R group and RVVC-S group (P0.05). CDR1,CDR2,MDR1 was expressed in all Candida albicans, only CDR1 gene expression was different: 5-fluorocytosine RVVC-R group was significantly higher than WC-R group (P0.05), miconazole was sensitive to miconazole. VVC-I R group was higher than VVC-S group (P0.05), fluconazole VVC-I group was higher than VVC-S group and VVC-R group (P0.05), but there was no difference between VVC-S group and WC-R group (P0.05). Conclusion: 1 the main pathogenic bacteria of recurrent vulvovaginal candidiasis are still Candida albicans; (2) the chromogenic medium was suitable for rapid identification of Candida albicans in clinic, but the accuracy of identification for non-Candida albicans was limited, the sensitivity of antifungal agents against Candida albicans caused by 3RVVC and VVC was generally the same, and nystatin was the first choice in the treatment of RVVC. 4PDR1 + CDR1 gene may be the drug-resistance related gene of Candida albicans in RWC and VVC patients, and besides the increase of drug resistance gene expression, there may be other more important related factors in the mechanism of drug resistance of RWC.
【學位授予單位】：昆明醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R711.31
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