Evaluation of nucleic acid sequencing of the D1/D2 region of the large subunit of the 28S rDNA and the internal transcribed spacer region using smartgene idn software for identification of filamentous fungi in a clinical laboratory

Nicole P. Kwiatkowski, Wisal M. Babiker, William G. Merz, Karen C. Carroll, Sean X. Zhang

Research output: Contribution to journalArticle

Abstract

Filamentous fungal infections have recently increased because of the increasing numbers of immunocompromised hosts. In this study, we evaluated DNA sequencing of the D1/D2 region of the large subunit of the 28S ribosomal RNA gene and the internal transcribed spacer (ITS) region using SmartGene (SG; SmartGene Inc., Raleigh, NC) for the identification of a broad range of commonly encountered filamentous fungi. The SG proofreaders were used to upload, align, and edit fragments, and the resultant sequences were interpreted using the quality-controlled SG database. The results were compared with reference identifications using conventional phenotypic methods or ITS DNA sequences obtained from GenBank if phenotypic identifications were inconclusive. A total of 146 clinical isolates were included in this study, representing 49 different genera. The overall agreements of the D1/D2 and the ITS sequencing methods to reference identification were 97.2% (95% CI, 93.1% to 98.9%) and 97.7% (95% CI, 92.8% to 99.4%), respectively. Of the 146 isolates, 18 (12.3%) did not amplify using the ITS universal primers after repeated attempts and, therefore, could not be sequenced using this target. Correct identification was achieved for 100% (95% CI, 97.4% to 100%) of the isolates when applying both the D1/D2 and ITS targets. In summary, DNA sequencing using SG software provides a rapid, accurate, and reliable tool for the identification of filamentous fungi in a clinical laboratory.

Original languageEnglish (US)
Pages (from-to)393-401
Number of pages9
JournalJournal of Molecular Diagnostics
Volume14
Issue number4
DOIs
StatePublished - Jul 1 2012

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ASJC Scopus subject areas

  • Pathology and Forensic Medicine
  • Molecular Medicine

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