title: Rapid identification and differentiation of fungal DNA in dermatological specimens by LightCycler PCR.
abstract: The aim was to develop a LightCycler PCR method for the rapid detection and differentiation of fungal DNA in dermatological specimens such as skin scales and skin swabs. LightCycler PCR assays were established for seven primer sets specific for fungal DNA. For each primer set LightCycler melting points were defined by amplification of DNA from 21 fungi and sensitivity was determined by amplification of serial dilutions of fungal DNA. A protocol was established that allows detection and differentiation of mould and yeast DNA with one highly sensitive PCR reaction by assessment of LightCycler melting points. Two subsequent LightCycler PCR reactions and one RFLP reaction allowed the differentiation of dermatophytes and non-dermatophyte moulds and the subclassification of yeasts. Analysis of clinical samples from 38 patients with fungal skin diseases provided conclusive new diagnostic information in 9/38 cases (23.7 %) by this PCR protocol that was not equally provided by direct microscopy and mycological culture. Thus the LightCycler PCR protocol established here represents a rapid diagnostic tool that aids in the diagnosis of fungal skin disease in a substantial number of patients.
authors.affiliation: Department of Dermatology and Allergology, Hannover Medical University, Ricklinger Str. 5, D-30449 Hannover, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Gutzmer
authors.affiliation: Department of Dermatology and Allergology, Hannover Medical University, Ricklinger Str. 5, D-30449 Hannover, Germany.
authors.forename: Susanne
authors.initials: S
authors.lastname: Mommert
authors.affiliation: Department of Dermatology and Allergology, Hannover Medical University, Ricklinger Str. 5, D-30449 Hannover, Germany.
authors.forename: Uta
authors.initials: U
authors.lastname: Küttler
authors.affiliation: Department of Dermatology and Allergology, Hannover Medical University, Ricklinger Str. 5, D-30449 Hannover, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Werfel
authors.affiliation: Department of Dermatology and Allergology, Hannover Medical University, Ricklinger Str. 5, D-30449 Hannover, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Kapp
mesh_terms: D004271:DNA, Fungal / Q000302:isolation & purification*; D003881:Dermatomycoses / Q000175:diagnosis / Q000382:microbiology*; D005658:Fungi / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D016533:Mycological Typing Techniques / Q000379:methods; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D012867:Skin / Q000382:microbiology; D013696:Temperature
chemical_list: D004271:DNA, Fungal