title: Development and validation of a multiplex PCR for detection of Scedosporium spp. in respiratory tract specimens from patients with cystic fibrosis.
abstract: The emergence of Scedosporium infections in diverse groups of individuals, which are often treatment refractory, warrants timely and accurate laboratory diagnosis. Species- or group-specific primers based on internal transcribed spacer (ITS) sequence polymorphisms were designed for Scedosporium aurantiacum, Scedosporium dehoogii, Scedosporium prolificans, Pseudallescheria boydii species complex (former clade 5)/Pseudallescheria apiosperma (formerly classified as S. apiospermum sensu lato) and Pseudallescheria minutispora. Primers for S. aurantiacum, S. prolificans, and P. boydii species complex/P. apiosperma were incorporated into a multiplex PCR assay for the detection and identification of the three major clinically important Scedosporium species and validated using sputum specimens collected from patients seen at a major Australian cystic fibrosis clinic. The multiplex PCR assay showed 100% specificity in identifying the three major clinically relevant Scedosporium species from pure culture. When evaluated using DNA extracts from sputa, sensitivity and specificity of the multiplex PCR assay were 62.1% and 97.2%, respectively. This highly species-specific multiplex PCR assay offers a rapid and simple method of detection of the most clinically important Scedosporium species in respiratory tract specimens.
authors.affiliation: Molecular Mycology Research Laboratory, Medical School-Westmead Hospital, Westmead, NSW 2145, Australia.
authors.forename: Azian
authors.initials: A
authors.lastname: Harun
authors.forename: Christopher C
authors.initials: CC
authors.lastname: Blyth
authors.forename: Felix
authors.initials: F
authors.lastname: Gilgado
authors.forename: Peter
authors.initials: P
authors.lastname: Middleton
authors.forename: Sharon C-A
authors.initials: SC
authors.lastname: Chen
authors.forename: Wieland
authors.initials: W
authors.lastname: Meyer
mesh_terms: D003550:Cystic Fibrosis / Q000150:complications*; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000235:genetics; D006801:Humans; D008172:Lung Diseases, Fungal / Q000175:diagnosis* / Q000382:microbiology; D008271:Mycetoma / Q000175:diagnosis* / Q000382:microbiology; D016133:Polymerase Chain Reaction / Q000379:methods*; D021681:Scedosporium / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
chemical_list: D017931:DNA Primers; D004271:DNA, Fungal; D021903:DNA, Ribosomal Spacer