title: MALDI-TOF MS proteomic phenotyping of filamentous and other fungi from clinical origin.
abstract: 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 "proteomic phenotypes". A growth time-course at 30°C on Sabouraud agar medium was performed for the 22 "phenotypes" at 48, 72, 96 and 120h 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 120h 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.
authors.affiliation: Parasitology Unit, Department of Laboratory Medicine, Bambino Gesù Children's Hospital, IRCCS, Piazza Sant'Onofrio 4, Rome, Italy.
authors.forename: Federica
authors.initials: F
authors.lastname: Del Chierico
authors.forename: Andrea
authors.initials: A
authors.lastname: Masotti
authors.forename: Manuela
authors.initials: M
authors.lastname: Onori
authors.forename: Ersilia
authors.initials: E
authors.lastname: Fiscarelli
authors.forename: Livia
authors.initials: L
authors.lastname: Mancinelli
authors.forename: Gabriella
authors.initials: G
authors.lastname: Ricciotti
authors.forename: Federico
authors.initials: F
authors.lastname: Alghisi
authors.forename: Leopoldo
authors.initials: L
authors.lastname: Dimiziani
authors.forename: Cesare
authors.initials: C
authors.lastname: Manetti
authors.forename: Andrea
authors.initials: A
authors.lastname: Urbani
authors.forename: Maurizio
authors.initials: M
authors.lastname: Muraca
authors.forename: Lorenza
authors.initials: L
authors.lastname: Putignani
mesh_terms: D000465:Algorithms; D005656:Fungal Proteins / Q000378:metabolism*; D003904:Mitosporic Fungi / Q000302:isolation & purification / Q000378:metabolism*; D009181:Mycoses / Q000175:diagnosis / Q000378:metabolism / Q000382:microbiology; D010449:Peptide Mapping / Q000379:methods*; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D005656:Fungal Proteins