本文選題：MALDI-TOF + MS　； 參考：《青島大學(xué)》2017年碩士論文

【摘要】：目的評(píng)價(jià)基質(zhì)輔助激光解吸電離飛行時(shí)間質(zhì)譜技術(shù)(matrix-assisted laser desorption/ionization-time of flight mass spectrometry,MALDI-TOF MS)在真菌鑒定中的應(yīng)用價(jià)值。方法關(guān)于絲狀真菌鑒定應(yīng)用:收集2015年8月~2016年12月青島市多家醫(yī)院患者標(biāo)本中分離培養(yǎng)出的絲狀真菌63株,購(gòu)買(mǎi)5株標(biāo)準(zhǔn)菌株,篩選出MALDI-TOF MS用于絲狀真菌鑒定的最佳培養(yǎng)基、培養(yǎng)時(shí)間、培養(yǎng)方式;同時(shí)進(jìn)行MALDI-TOF MS鑒定、形態(tài)學(xué)鑒定,形態(tài)學(xué)不能鑒定的菌株進(jìn)行內(nèi)轉(zhuǎn)錄間隔ITS序列測(cè)定;對(duì)三種方法的結(jié)果進(jìn)行對(duì)比,以評(píng)價(jià)MALDI-TOF MS在絲狀真菌鑒定中的能力。關(guān)于念珠菌鑒定應(yīng)用:利用質(zhì)控菌株ATCC 90028白色念珠菌確定用于MALDI-TOF MS鑒定的最佳培養(yǎng)條件;然后,從我院2015年11月~2016年6月MALDI-TOF MS鑒定的299株念珠菌中隨機(jī)選取101株念珠菌,對(duì)上述念珠菌同時(shí)進(jìn)行MALDI-TOF MS鑒定、顯色培養(yǎng)鑒定、ATB Expression鑒定,對(duì)三種方法鑒定結(jié)果進(jìn)行對(duì)比分析,以評(píng)價(jià)MALDI-TOF MS在念珠菌鑒定中的能力。結(jié)果篩選MALDI-TOF MS用于絲狀真菌鑒定的最佳培養(yǎng)基為沙氏葡萄糖液體培養(yǎng)基,培養(yǎng)時(shí)間為24h。67株絲狀真菌MALDI-TOF MS鑒定結(jié)果與形態(tài)學(xué)鑒定和分子生物學(xué)鑒定結(jié)果基本一致,在屬、種水平的鑒定總一致率分別為98.5%、94.1%,對(duì)曲霉菌屬鑒定種水平一致率達(dá)到100%。MALDI-TOF MS對(duì)101株念珠菌進(jìn)行鑒定結(jié)果與顯色培養(yǎng)和ATB Expression鑒定結(jié)果比較,在屬水平鑒定一致率為98.02%,在種水平鑒定一致率為97.03%。結(jié)論MALDI-TOF MS技術(shù)在絲狀真菌以及念珠菌鑒定中符合率較高,相比于傳統(tǒng)方法操作簡(jiǎn)單、成本低、高通量、耗時(shí)短,應(yīng)用于臨床微生物實(shí)驗(yàn)室真菌的常規(guī)鑒定可以極大提高檢測(cè)效率。但是,真菌蛋白質(zhì)的提取量對(duì)于檢測(cè)結(jié)果影響顯著,分析前的樣品處理標(biāo)準(zhǔn)化十分重要。
[Abstract]:Objective to evaluate the application value of matrix-assisted laser desorption/ionization-time of flight mass spectrometric MALDI-TOF MS in fungal identification. Methods for the identification of filamentous fungi, 63 strains of filamentous fungi were collected from patients in Qingdao from August 2015 to December 2016, 5 standard strains were purchased and the best medium for identification of filamentous fungi by MALDI-TOF MS was selected. Culture time, culture method, MALDI-TOF MS identification, morphological identification, morphological identification, ITS sequence analysis of internal transcription interval were performed. The results of the three methods were compared. To evaluate the ability of MALDI-TOF MS in the identification of filamentous fungi. On the application of Candida candida identification: the best culture conditions for MALDI-TOF MS identification were determined by ATCC 90028 strain, and then, 101 Candida albicans were randomly selected from MALDI-TOF MS identified in our hospital from November 2015 to June 2016. At the same time, the Candida species were identified by MALDI-TOF MS, and the results of the three methods were compared and analyzed in order to evaluate the ability of MALDI-TOF MS in the identification of Candida. Results the best medium for identification of filamentous fungi by MALDI-TOF MS was glucose liquid medium. The results of MALDI-TOF MS identification of filamentous fungi of 24h.67 strain were consistent with those of morphology and molecular biology. The identification rate of aspergillus was 98.5% and 94.1%, respectively. The identification rate of Aspergillus was up to that of 100%.MALDI-TOF MS. The results of identification of 101 Candida strains were compared with those of chromogenic culture and ATB Expression. The consistent rate of identification was 98.02 at the genus level and 97.03 at the species level. Conclusion the coincidence rate of MALDI-TOF MS in the identification of filamentous fungi and candida is higher than that of traditional methods, which has the advantages of simple operation, low cost, high throughput and short time consuming. The routine identification of fungi in clinical microorganism laboratory can greatly improve the detection efficiency. However, the extraction amount of fungal protein has a significant effect on the detection results, and the standardization of sample treatment before analysis is very important.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R446.5

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