Evaluation of MALDI-TOF MS, sequencing of D2 LSU rRNA and internal transcribed spacer regions (ITS) for the identification of filamentous fungi isolated from a pharmaceutical facility

• Published in: Journal of pharmaceutical and biomedical analysis Vol. 234; p. 115531
• Main Authors: Moreira, Filipe Mercês, Pereira, Pamalla de Araujo, Miranda, Rebeca Vitória da Silva Lage de, Reis, Cristhiane Moura Falavina dos, Braga, Lygia Maria Paulo da Silva, de Andrade, Joyce Modesto, do Nascimento, Luciane Gomes, Mattoso, Josiane Machado Vieira, Forsythe, Stephen James, da Costa, Luciana Veloso, Brandão, Marcelo Luiz Lima
• Format: Journal Article
• Language: English
• Published: England Elsevier B.V 20-09-2023
• Subjects: Fungi filamentous; Pharmaceutical facility; Protein analysis; Sequencing; Pharmaceutical facility; Protein analysis; Sequencing; Fungi filamentous
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/j.jpba.2023.115531

The identification of filamentous fungi through culture characterization may be hampered by phenotypic variability. Information obtained from the identification of microorganisms are important for investigation of sources of contamination of a product or process. The aim of this study was to identify filamentous fungal strains (n = 50) isolated from a pharmaceutical facility by using Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), as well as D2 domain of the large-subunit (LSU) ribosomal RNA gene and internal transcribed spacer regions (ITS) sequencing. MALDI-TOF MS system only identified five strains at the species level, while 45 were not identified. The analysis through GenBank allowed the identification of up to 19 strains at the species level, while MycoBank allowed the identification of up to nine strains at the species level. The databases identified up to 11 genera: Penicillium, Aspergillus, Cladosporium, Chaetomium, Coniochaeta, Curvularia, Diaporthe, Fusarium, Trichoderma, Rhizopus and Microdochium. MALDI-TOF MS showed an insufficient database to identify the species of fungi. DNA sequencing was the best methodology to identify to the genus level but was unable to differentiate between closely related species. Therefore further methods for the identification of filamentous fungi from pharmaceutical areas at species level need to be developed.
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•MALDI-TOF MS database was insufficient to identify environmental filamentous fungi.•The ITS region identified 19 filamentous fungi strains at the species level.•D2 LSU rDNA showed highest capacity to identify filamentous fungi at genus level.•Closely related species could not be differentiated with D2 LSU rRNA and ITS.•Coniochaeta, Diaporthe and Microdochium were found in a pharmaceutical facility.