title: Molecular identification of unusual pathogenic yeast isolates by large ribosomal subunit gene sequencing: 2 years of experience at the United kingdom mycology reference laboratory.
abstract: Rapid identification of yeast isolates from clinical samples is particularly important given their innately variable antifungal susceptibility profiles. We present here an analysis of the utility of PCR amplification and sequence analysis of the hypervariable D1/D2 region of the 26S rRNA gene for the identification of yeast species submitted to the United Kingdom Mycology Reference Laboratory over a 2-year period. A total of 3,033 clinical isolates were received from 2004 to 2006 encompassing 50 different yeast species. While more than 90% of the isolates, corresponding to the most common Candida species, could be identified by using the AUXACOLOR2 yeast identification kit, 153 isolates (5%), comprised of 47 species, could not be identified by using this system and were subjected to molecular identification via 26S rRNA gene sequencing. These isolates included some common species that exhibited atypical biochemical and phenotypic profiles and also many rarer yeast species that are infrequently encountered in the clinical setting. All 47 species requiring molecular identification were unambiguously identified on the basis of D1/D2 sequences, and the molecular identities correlated well with the observed biochemical profiles of the various organisms. Together, our data underscore the utility of molecular techniques as a reference adjunct to conventional methods of yeast identification. Further, we show that PCR amplification and sequencing of the D1/D2 region reliably identifies more than 45 species of clinically significant yeasts and can also potentially identify new pathogenic yeast species.
authors.affiliation: Mycology Reference Laboratory, Health Protection Agency South-West Regional Laboratory, Myrtle Road, Bristol BS2 8EL, United Kingdom. Chris.Linton@bris.ac.uk
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Linton
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Borman
authors.forename: Grace
authors.initials: G
authors.lastname: Cheung
authors.forename: Ann D
authors.initials: AD
authors.lastname: Holmes
authors.forename: Adrien
authors.initials: A
authors.lastname: Szekely
authors.forename: Michael D
authors.initials: MD
authors.lastname: Palmer
authors.forename: Paul D
authors.initials: PD
authors.lastname: Bridge
authors.forename: Colin K
authors.initials: CK
authors.lastname: Campbell
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Johnson
mesh_terms: D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D004275:DNA, Ribosomal / Q000737:chemistry / Q000235:genetics; D005838:Genotype; D006801:Humans; D016533:Mycological Typing Techniques; D009181:Mycoses / Q000175:diagnosis / Q000382:microbiology*; D010641:Phenotype; D016133:Polymerase Chain Reaction; D012335:RNA, Ribosomal / Q000235:genetics*; D017422:Sequence Analysis, DNA; D006113:United Kingdom; D015003:Yeasts / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification
chemical_list: D004271:DNA, Fungal; D004275:DNA, Ribosomal; D012335:RNA, Ribosomal; C065096:RNA, ribosomal, 26S