MALDI-TOF MS在真菌鉴定中的应用研究

发布时间：2018-04-29 15:12

本文选题：MALDI-TOF + MS　；参考：《青岛大学》2017年硕士论文

【摘要】：目的评价基质辅助激光解吸电离飞行时间质谱技术(matrix-assisted laser desorption/ionization-time of flight mass spectrometry,MALDI-TOF MS)在真菌鉴定中的应用价值。方法关于丝状真菌鉴定应用:收集2015年8月~2016年12月青岛市多家医院患者标本中分离培养出的丝状真菌63株,购买5株标准菌株,筛选出MALDI-TOF MS用于丝状真菌鉴定的最佳培养基、培养时间、培养方式;同时进行MALDI-TOF MS鉴定、形态学鉴定,形态学不能鉴定的菌株进行内转录间隔ITS序列测定;对三种方法的结果进行对比,以评价MALDI-TOF MS在丝状真菌鉴定中的能力。关于念珠菌鉴定应用:利用质控菌株ATCC 90028白色念珠菌确定用于MALDI-TOF MS鉴定的最佳培养条件;然后,从我院2015年11月~2016年6月MALDI-TOF MS鉴定的299株念珠菌中随机选取101株念珠菌,对上述念珠菌同时进行MALDI-TOF MS鉴定、显色培养鉴定、ATB Expression鉴定,对三种方法鉴定结果进行对比分析,以评价MALDI-TOF MS在念珠菌鉴定中的能力。结果筛选MALDI-TOF MS用于丝状真菌鉴定的最佳培养基为沙氏葡萄糖液体培养基,培养时间为24h。67株丝状真菌MALDI-TOF MS鉴定结果与形态学鉴定和分子生物学鉴定结果基本一致,在属、种水平的鉴定总一致率分别为98.5%、94.1%,对曲霉菌属鉴定种水平一致率达到100%。MALDI-TOF MS对101株念珠菌进行鉴定结果与显色培养和ATB Expression鉴定结果比较,在属水平鉴定一致率为98.02%,在种水平鉴定一致率为97.03%。结论MALDI-TOF MS技术在丝状真菌以及念珠菌鉴定中符合率较高,相比于传统方法操作简单、成本低、高通量、耗时短,应用于临床微生物实验室真菌的常规鉴定可以极大提高检测效率。但是,真菌蛋白质的提取量对于检测结果影响显著,分析前的样品处理标准化十分重要。
[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.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R446.5

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