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Journal of Clinical Microbiology, February 2001, p. 685-690, Vol. 39, No. 2
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.2.685-690.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Rapid Discrimination among Dermatophytes, Scytalidium spp., and Other Fungi with a PCR-Restriction Fragment Length Polymorphism Ribotyping Method

Marie Machouart-Dubach,1,* Claire Lacroix,1,2 Martine Feuilhade de Chauvin,1,2 Isabelle Le Gall,3 Catherine Giudicelli,3 Frédéric Lorenzo,4 and Francis Derouin1,2

Laboratoire de Parasitologie-Mycologie, UFR Lariboisière Saint-Louis-Université Paris 7, Faculté de Médecine, 75006 Paris,1 Laboratoire de Parasitologie-Mycologie, Hôpital Saint-Louis,2 and Centre d'Etude du Polymorphisme Humain, Hôpital Saint-Louis,3 75475 Paris Cedex 10, and Association HEMERA, 94424 Le Plessis-Trevise,4 France

Received 23 June 2000/Returned for modification 31 August 2000/Accepted 13 November 2000

Dermatomycoses are very common infections caused mainly by dermatophytes. Scytalidiosis is a differential mycological diagnosis, especially in tropical and subtropical areas. Since a culture-based diagnosis takes 2 to 3 weeks, we set up a PCR-restriction fragment length polymorphism (RFLP) method for rapid discrimination of these fungi in clinical samples. The hypervariable V4 domain of the small ribosomal subunit 18S gene was chosen as the target for PCR. The corresponding sequences from 19 fungal species (9 dermatophytes, 2 Scytalidium species, 6 other filamentous fungi, and 2 yeasts) were obtained from databases or were determined in the laboratory. Sequences were aligned to design primers for dermatophyte-specific PCR and to identify digestion sites for RFLP analysis. The reliability of PCR-RFLP for the diagnosis of dermatomycosis was assessed on fungal cultures and on specimens from patients with suspected dermatomycosis. Two sets of primers preferentially amplified fungal DNA from dermatophytes (DH1L and DH1R) or from Scytalidium spp. (DH2L and DH1R) relative to DNA from bacteria, yeasts, some other filamentous fungi, and humans. Digestion of PCR products with EaeI or BamHI discriminated between dermatophytes and Scytalidium species, as shown with cultures of 31 different fungal species. When clinical samples were tested by PCR-RFLP, blindly to mycological findings, the results of the two methods agreed for 74 of 75 samples. Dermatophytes and Scytalidium spp. can thus be readily discriminated by PCR-RFLP within 24 h. This method can be applied to clinical samples and is suited to rapid etiologic diagnosis and treatment selection for patients with dermatomycosis.

* Corresponding author. Mailing address: Laboratoire de Parasitologie-Mycologie, UFR Lariboisière Saint-Louis-Université Paris 7, Faculté de Médecine, 15 Rue de l'École de Médecine, 75006 Paris, France. Phone: 33-1-43-29-65-25. Fax: 33-1-43-29-51-92. E-mail: paracord{at}wanadoo.fr.

Journal of Clinical Microbiology, February 2001, p. 685-690, Vol. 39, No. 2
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.2.685-690.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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