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Journal of Clinical Microbiology, February 2007, p. 380-385, Vol. 45, No. 2
0095-1137/07/$08.00+0     doi:10.1128/JCM.01862-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Development and Clinical Application of a Panfungal PCR Assay To Detect and Identify Fungal DNA in Tissue Specimens{triangledown}

Anna Lau,1 Sharon Chen,2,3 Tania Sorrell,1,2 Dee Carter,4 Richard Malik,5,6 Patricia Martin,6 and Catriona Halliday2,3*

Faculty of Medicine,1 Centre for Infectious Diseases and Microbiology, Westmead Millennium Institute,2 School of Molecular and Microbial Biosciences,4 Post Graduate Foundation in Veterinary Science,5 Veterinary Pathology Diagnostic Services, Faculty of Veterinary Science, University of Sydney,6 Centre for Infectious Diseases and Microbiology Laboratory Services, Institute of Clinical Pathology and Medical Research, Sydney West Area Health Service, Sydney, New South Wales, Australia3

Received 8 September 2006/ Returned for modification 7 November 2006/ Accepted 15 November 2006

Given the rise in the incidence of invasive fungal infections (IFIs) and the expanding spectrum of fungal pathogens, early and accurate identification of the causative pathogen is essential. We developed a panfungal PCR assay that targets the internal transcribed spacer 1 (ITS1) region of the ribosomal DNA gene cluster to detect fungal DNA in fresh and formalin-fixed, paraffin-embedded (PE) tissue specimens from patients with culture-proven (n = 38) or solely histologically proven (n = 24) IFIs. PCR products were sequenced and compared with sequences in the GenBank database to identify the causal pathogen. The molecular identification was correlated with results from histological examination and culture. The assay successfully detected and identified the fungal pathogen in 93.6% and 64.3% of culture-proven and solely histologically proven cases of IFI, respectively. A diverse range of fungal genera were identified, including species of Candida, Cryptococcus, Trichosporon, Aspergillus, Fusarium, Scedosporium, Exophiala, Exserohilum, Apophysomyces, Actinomucor, and Rhizopus. For five specimens, molecular analysis identified a pathogen closely related to that identified by culture. All PCR-negative specimens (n = 10) were PE tissues in which fungal hyphae were visualized. The results support the use of the panfungal PCR assay in combination with conventional laboratory tests for accurate identification of fungi in tissue specimens.

* Corresponding author. Mailing address: Centre for Infectious Diseases and Microbiology, Westmead Hospital, Westmead, New South Wales, 2145, Australia. Phone: 61-2-9845 6255. Fax: 61-2-9893 8659. E-mail: catrionah{at}icpmr.wsahs.nsw.gov.au.

{triangledown} Published ahead of print on 22 November 2006.

Journal of Clinical Microbiology, February 2007, p. 380-385, Vol. 45, No. 2
0095-1137/07/$08.00+0     doi:10.1128/JCM.01862-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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