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TitleMultiplex PCR based strategy for detection of fungal pathogen DNA in patients with suspected invasive fungal infections
Author(s)Carvalho-Pereira, Joana
Fernandes, Filipa
Araújo, Ricardo
Springer, Jan
Loeffler, Juergen
Buitrago, María José
Pais, Célia
Sampaio, Paula
Keywordsmolecular diagnosis
fungal infections
multiplex PCR
Candida sp
Aspergillus sp
Rhizopus arrhizus
Issue date2020
PublisherMultidisciplinary Digital Publishing Institute (MDPI)
JournalJournal of Fungi
CitationCarvalho-Pereira, J.; Fernandes, F.; Araújo, R.; Springer, J.; Loeffler, J.; Buitrago, M.J.; Pais, C.; Sampaio, P. Multiplex PCR Based Strategy for Detection of Fungal Pathogen DNA in Patients with Suspected Invasive Fungal Infections. J. Fungi 2020, 6, 308.
Abstract(s)A new and easy polymerase chain reaction (PCR) multiplex strategy, for the identification of the most common fungal species involved in invasive fungal infections (IFI) was developed in this work. Two panels with species-specific markers were designed, the <i>Candida Panel</i> for the identification of <i>Candida</i> species, and the <i>Filamentous Fungi Panel</i> for the identification of <i>Aspergillus</i> species and <i>Rhizopus</i><i>arrhizus</i>. The method allowed the correct identification of all targeted pathogens using extracted DNA or by colony PCR, showed no cross-reactivity with nontargeted species and allowed identification of different species in mixed infections. Sensitivity reached 10 to 1 pg of DNA and was suitable for clinical samples from sterile sites, with a sensitivity of 89% and specificity of 100%. Overall, the study showed that the new method is suitable for the identification of the ten most important fungal species involved in IFI, not only from positive blood cultures but also from clinical samples from sterile sites. The method provides a unique characteristic, of seeing the peak in the specific region of the panel with the correct fluorescence dye, that aids the ruling out of unspecific amplifications. Furthermore, the panels can be further customized, selecting markers for different species and/or resistance genes.
Publisher version
AccessOpen access
Appears in Collections:CBMA - Artigos/Papers

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