MALDI-TOF MS proteomic phenotyping of filamentous and other fungi from clinical origin

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• 作者：Federica Del Chierico^a ; Andrea Masotti^b ; Manuela Onori^c ; Ersilia Fiscarelli^d ; Livia Mancinelli^c ; Gabriella Ricciotti^e ; Federico Alghisi^f ; Leopoldo Dimiziani^g ; Cesare Manetti^h ; Andrea Urbaniⁱ ; ^j ; Maurizio Muraca^d ; Lorenza Putignani^a ; ^{lorenza.putignani@opbg.net}
• 关键词：Emerging/uncommon pathogenic fungi ; Cystic fibrosis (CF) patients ; MALDI-TOF MS ; Solid medium-based-customized database ; Fingerprinting classifiers ; MS-based analytical identification (ID) assay
• 刊名：Journal of Proteomics
• 出版年：2012
• 期刊代码：P37_18743919
• 类别：ch
• 出版时间：18 June, 2012
• 卷：75
• 期：11
• 页码：3314-3330
• 文件大小：2371 K

摘要

Major changes in medical, intensive care and organ transplantation practices are drastically increasing the risk of fungal opportunistic infections. We designed and set-up a MALDI-TOF MS-based assay to identify the most isolated and emerging therapy-refractory/uncommon fungi from cystic fibrosis (CF) and immunocompromised patients.

Two-hundred and thirty isolates from 10 different genera (Aspergillus, Emericella, Fusarium, Geosmithia, Neosartorya, Penicillium, Pseudallescheria, Scedosporium, Talaromyces, Fomitopsis), investigated during routine diagnostic efforts, were correlated to 22 laboratory-adapted reference MALDI-TOF MS 鈥減roteomic phenotypes鈥? A growth time-course at 30 掳C on Sabouraud agar medium was performed for the 22 鈥減henotypes鈥?at 48, 72, 96 and 120 h points. The best peptide extraction conditions for full recovery of conidia- or asci-producing multihyphal morph structures and the highest intra- and inter-class profiling correlation were identified for the 120 h point spectra dataset, from which an engineered library derived (pre-analytical phase). Fingerprinting classifiers, selected by Wilcoxon/Kruskal-Wallis algorithm, were computed by Genetic Algorithm, Support Vector Machine, Supervised Neuronal Network and Quick Classifier model construction. MS identification (ID) of clinical isolates was referred to genotyping (GT) and, retrospectively, compared to routine morphotyping (MT) IDs (analytical phase).

Proteomic phenotyping is revolutionizing diagnostic mycology as fully reflecting species/morph varieties but often overcoming taxonomic hindrance.