本文選題：白念珠菌 + Mrr2��； 參考：《山西醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的:1、了解本地VVC患者念珠菌感染的菌種分布情況及其對FCA的敏感性;2、探討VVC患者臨床分離白念珠菌Mrr2突變與FCA耐藥的關(guān)系;3、探討VVC患者臨床分離白念珠菌Mrr2、CDR1的mRNA表達(dá)水平與FCA耐藥的關(guān)系,并進(jìn)一步分析Mrr2與CDR1表達(dá)水平在FCA耐藥中的相互調(diào)控作用;4、探討VVC患者臨床分離白念珠菌Mrr2基因突變與Mrr2的mRNA表達(dá)水平是否對FCA耐藥具有相互作用。方法:采集2015年11月至2016年5月山西醫(yī)科大學(xué)第二醫(yī)院皮膚性病科就診的可疑VVC患者陰道分泌物標(biāo)本155份,并經(jīng)過真菌鏡檢、CHROMagar念珠菌顯色培養(yǎng)基及API 20C AUX念珠菌鑒定系統(tǒng)進(jìn)行菌種鑒定,獲得102株念珠菌。對其中的80株白念珠菌應(yīng)用M27-A3微量肉湯稀釋方法進(jìn)行FCA藥物敏感性試驗。對80株白念珠菌分別提取DNA、聚合酶鏈反應(yīng)(Polymerase Chain Reaction,PCR)擴(kuò)增Mrr2基因片段、測序,尋找突變位點。對白念珠菌FCA耐藥組和敏感組菌株分別提取總RNA,逆轉(zhuǎn)錄合成cDNA,實時熒光定量PCR(Real-time Fluorescence Quantitative PCR,FQ-RT-PCR)檢測Mrr2、CDR1的mRNA表達(dá)量,并做相關(guān)統(tǒng)計學(xué)分析。結(jié)果:1、從155份陰道分泌物標(biāo)本中,共獲得念珠菌菌株102株。其中白念珠菌80株(78.4%),光滑念珠菌15株(14.7%),熱帶念珠菌4株(3.9%),近平滑念珠菌2株(2.0%),克柔念珠菌1株(1.0%)。2、對80株白念珠菌接種活化,并使用M27-A3微量肉湯稀釋法進(jìn)行FCA藥物敏感性試驗,結(jié)果顯示:80株白念珠菌中有40株對FCA敏感,10株對FCA呈劑量依賴性敏感,30株對FCA耐藥,耐藥率為37.5%。3、對80株白念珠菌的Mrr2基因進(jìn)行測序,其中FCA耐藥組Mrr2基因突變率為56.67%;FCA敏感組Mrr2基因突變率為26.08%。對白念珠菌Mrr2基因突變與FCA耐藥的關(guān)系采用四格表卡方檢驗,得出P值0.05,差異具有統(tǒng)計學(xué)意義。耐藥組Mrr2基因突變率高于敏感組Mrr2的突變率。4、檢測白念珠菌FCA敏感組和耐藥組的Mrr2及CDR1的mRNA表達(dá)水平,經(jīng)統(tǒng)計學(xué)分析:(1)白念珠菌FCA耐藥組Mrr2的mRNA表達(dá)水平高于敏感組Mrr2的mRNA表達(dá)水平(1.10±0.50 VS 0.52±0.45),P0.05,差異具有統(tǒng)計學(xué)意義。(2)白念珠菌FCA耐藥組CDR1的mRNA表達(dá)水平高于敏感組CDR1的mRNA表達(dá)水平(0.42±0.294 VS 0.25±0.289),P0.05,差異具有統(tǒng)計學(xué)意義。(3)CDR1與Mrr2呈正相關(guān),r=37.6%。(4)Mrr2突變與Mrr2表達(dá)的交互作用分析,發(fā)現(xiàn)Mrr2基因發(fā)生突變且高表達(dá)組的耐藥性是基因未發(fā)生突變且基因低表達(dá)組的耐藥性的47.50倍,說明Mrr2基因突變與高表達(dá)對FCA耐藥性的產(chǎn)生存在協(xié)同作用。結(jié)論:1、本地區(qū)VVC患者最主要的致病菌種是白念珠菌,對FCA耐藥率較高,臨床治療VVC應(yīng)依據(jù)藥敏試驗結(jié)果,選用敏感抗真菌藥物進(jìn)行精準(zhǔn)治療。2、VVC患者臨床分離白念珠菌FCA耐藥性可能與Mrr2突變有關(guān)。3、CDR1高表達(dá)可能直接導(dǎo)致白念珠菌對FCA耐藥;Mrr2高表達(dá)可能直接導(dǎo)致白念珠菌對FCA耐藥,且Mrr2高表達(dá)還可促進(jìn)CDR1高表達(dá)進(jìn)而介導(dǎo)白念珠菌對FCA耐藥。4、Mrr2基因突變與高表達(dá)對FCA耐藥性的產(chǎn)生可能存在協(xié)同作用。
[Abstract]:Objective to investigate the distribution of Candida albicans in local VVC patients and their sensitivity to FCA. To explore the relationship between Mrr2 mutation of clinical isolates of Candida albicans and drug resistance to FCA in patients with VVC. The relationship between the expression of mRNA and the drug resistance of FCA. Furthermore, the mutual regulation of Mrr2 and CDR1 expression level in FCA resistance was analyzed. The relationship between Mrr2 gene mutation and Mrr2 mRNA expression level in VVC patients was studied to determine whether the Mrr2 gene mutation and Mrr2 mRNA expression level had interaction with FCA resistance. Methods: from November 2015 to May 2016, 155 specimens of vaginal secretions from suspected VVC patients in Department of Dermatology and venereal Diseases, second Hospital of Shanxi Medical University, were collected. The strain of Candida albicans was identified by API 20C AUX system and the culture medium of CHROMagar was used to identify the species of Candida albicans, and 102 strains of Candida were obtained. The susceptibility of 80 strains of Candida albicans to FCA was tested by M27-A3 broth dilution method. DNA was extracted from 80 strains of Candida albicans. The Mrr2 gene fragment was amplified by polymerase chain reaction (PCR) and sequenced to find the mutation site. The total RNAs were extracted from Candida albicans FCA resistant and susceptible strains respectively, and the cDNAs were synthesized by reverse transcription. The mRNA expression of MRR2 CDR1 was detected by real-time quantitative PCR(Real-time Fluorescence Quantitative PCR FQ-RT-PCR1, and the correlation statistical analysis was made. Results 102 strains of Candida were obtained from 155 vaginal secretions. Among them, 80 strains of Candida albicans were inoculated and activated, 15 strains of Candida smooth were inoculated and activated, 4 strains of Candida tropicalis were found to be 3. 9%, 2 strains were close to Candida smoothing, 2 strains were close to Candida smoothing, 1 strain was 1 strain of Candida korgii, and inoculated and activated to 80 strains of Candida albicans, and the sensitivity of FCA was tested by M27-A3 broth dilution method. The results showed that 40 of the 80 strains of Candida albicans were sensitive to FCA. 10 strains were dose-dependent sensitive to FCA. 30 strains were resistant to FCA. The drug resistance rate was 37.5%. The Mrr2 gene of 80 strains of Candida albicans was sequenced. The mutation rate of Mrr2 gene in FCA resistant group was 56.67 and 26.08 in FCA-sensitive group. The relationship between Mrr2 gene mutation of Candida albicans and drug resistance of FCA was tested by four-grid table-chi-square test, P value was 0.05, the difference was statistically significant. The mutation rate of Mrr2 gene in resistant group was higher than that in sensitive group. The mRNA expression of Mrr2 and CDR1 in susceptible and resistant group of Candida albicans FCA was detected. The mRNA expression level of Mrr2 in Candida albicans FCA resistant group was 1.10 鹵0.50 vs 0.52 鹵0.45 P 0.05, the difference was statistically significant. 2) the CDR1 mRNA expression level of Candida albicans FCA resistant group was higher than that of sensitive CDR1 group. The expression level was 0.42 鹵0.294 vs 0.25 鹵0.289 P0.05, the difference was statistically significant. There was a positive correlation between CDR1 and Mrr2. There was a positive correlation between the mutation of MRR2 and the expression of Mrr2. It was found that the drug resistance of Mrr2 gene mutation and high expression group was 47.50 times higher than that of non-mutation gene and low gene expression group, which indicated that there was a synergistic effect of Mrr2 gene mutation and high expression on the drug resistance of FCA. Conclusion the main pathogen of VVC patients in this area is Candida albicans, and the rate of drug resistance to FCA is high. The clinical treatment of VVC should be based on the results of drug sensitivity test. The drug resistance of Candida albicans isolated from clinical isolates of Candida albicans FCA may be related to the mutation of Mrr2. The high expression of CDR1 may directly lead to the high expression of MRR2 in Candida albicans resistant to FCA, which may lead to the resistance of Candida albicans to FCA, and the drug resistance of Candida albicans to FCA may be directly related to the drug resistance of Candida albicans to FCA. Moreover, the high expression of Mrr2 can promote the high expression of CDR1 and mediate the mutation of MRR2 gene in Candida albicans resistant to FCA and the production of drug resistance to FCA by high expression.
【學(xué)位授予單位】：山西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R446.5;R711.3

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