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DNA Microarray Based on Arrayed-Primer Extension Technique for Identification of Pathogenic Fungi Responsible for Invasive and Superficial Mycoses

Daniele Campa,^1, Arianna Tavanti,^2, Federica Gemignani,¹ Crocifissa S. Mogavero,² Ilaria Bellini,¹ Fabio Bottari,¹ Roberto Barale,¹ Stefano Landi,^1, and Sonia Senesi^2,*

Dipartimento di Biologia, Sezione di Genetica, Via Derna 1,¹ Sezione di Microbiologia, Via San Zeno 35-39, Università di Pisa, 56126 Pisa, Italy²

Received 12 July 2007/ Returned for modification 4 October 2007/ Accepted 14 December 2007

An oligonucleotide microarray based on the arrayed-primer extension (APEX) technique has been developed to simultaneously identify pathogenic fungi frequently isolated from invasive and superficial infections. Species-specific oligonucleotide probes complementary to the internal transcribed spacer 1 and 2 (ITS1 and ITS2) region were designed for 24 species belonging to 10 genera, including Candida species (Candida albicans, Candida dubliniensis, Candida famata, Candida glabrata, Candida tropicalis, Candida kefyr, Candida krusei, Candida guilliermondii, Candida lusitaniae, Candida metapsilosis, Candida orthopsilosis, Candida parapsilosis, and Candida pulcherrima), Cryptococcus neoformans, Aspergillus species (Aspergillus fumigatus and Aspergillus terreus), Trichophyton species (Trichophyton rubrum and Trichophyton tonsurans), Trichosporon cutaneum, Epidermophyton floccosum, Fusarium solani, Microsporum canis, Penicillium marneffei, and Saccharomyces cerevisiae. The microarray was tested for its specificity with a panel of reference and blinded clinical isolates. The APEX technique was proven to be highly discriminative, leading to unequivocal identification of each species, including the highly related ones C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Because of the satisfactory basic performance traits obtained, such as reproducibility, specificity, and unambiguous interpretation of the results, this new system represents a reliable method of potential use in clinical laboratories for parallel one-shot detection and identification of the most common pathogenic fungi.

Published ahead of print on 26 December 2007.

D.C., A.T., S.L., and S.S. contributed equally to this study.