title: Utility of DNA sequencing for direct identification of invasive fungi from fresh and formalin-fixed specimens.
abstract: OBJECTIVE
  To describe and discuss the utility and potential pitfalls of ribosomal RNA locus sequencing for direct identification of invasive fungi from fresh and formalin-fixed, paraffin-embedded specimens.
  
  
  METHODS
  DNA was extracted from fresh and formalin-fixed, paraffin-embedded tissue and subjected to real-time polymerase chain reaction (PCR) targeting ITS2 and D2 regions of fungal ribosomal RNA locus. Cycle sequencing was performed on PCR products, and the identity of sequences was determined using a public database.
  
  
  RESULTS
  Four clinical cases of invasive fungal infection are presented to illustrate the utility of DNA sequencing for determining etiology when microbiological culture is negative, for shortening the time to identification of slow-growing fungi, for guiding antifungal therapy, and for shedding light on the pathogenesis of disseminated fungal infection.
  
  
  CONCLUSIONS
  Fungal ribosomal RNA locus sequencing from fresh or formalin-fixed, paraffin-embedded specimens is a powerful tool for rapid and accurate diagnosis of patients with culture-negative or uncultured invasive mycosis.
authors.affiliation: Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, CA, USA.
authors.forename: Pablo A
authors.initials: PA
authors.lastname: Moncada
authors.forename: Indre
authors.initials: I
authors.lastname: Budvytiene
authors.forename: Dora Y
authors.initials: DY
authors.lastname: Ho
authors.forename: Stanley C
authors.initials: SC
authors.lastname: Deresinski
authors.forename: Jose G
authors.initials: JG
authors.lastname: Montoya
authors.forename: Niaz
authors.initials: N
authors.lastname: Banaei
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D004271:DNA, Fungal / Q000235:genetics*; D005557:Formaldehyde; D005658:Fungi / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D008297:Male; D009181:Mycoses / Q000175:diagnosis* / Q000235:genetics / Q000378:metabolism; D016612:Paraffin Embedding; D017422:Sequence Analysis, DNA; D016707:Tissue Fixation
chemical_list: D004271:DNA, Fungal; D005557:Formaldehyde
keywords: Fungi; Identification; Invasive; Paraffin-embedded; Sequencing