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Multiplex PCR assay for the detection of common dermatophyte nail infections
Author(s) -
Dhib I.,
Fathallah A.,
Yaacoub A.,
Hadj Slama F.,
Said M. B.,
Zemni R.
Publication year - 2014
Publication title -
mycoses
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.13
H-Index - 69
eISSN - 1439-0507
pISSN - 0933-7407
DOI - 10.1111/myc.12096
Subject(s) - dermatophyte , trichophyton rubrum , dna extraction , multiplex polymerase chain reaction , nail (fastener) , trichophyton , polymerase chain reaction , direct examination , biology , internal transcribed spacer , microbiology and biotechnology , gene , genetics , ribosomal rna , antifungal , materials science , law , political science , metallurgy
Summary Onychomycosis is one of the most prevalent dermatophytic diseases. Mycological methods used in the conventional diagnosis may not be optimal. Multiplex ( MX ) PCR was reported as a reliable alternative. Dermatophyte gene sequence records were used to design a MX PCR for detection and identification of dermatophytes in nail specimens. A MX PCR method based on the amplification of the chitin synthase 1 and internal transcribed spacer genes was developed. The study included 93 strains of dermatophytes and non‐dermatophytic fungi, six dermatophytic reference strains and 201 nail specimens from patients with dermatophytic onyxis. DNA extraction directly from nail samples was carried out by using the QIAamp DNA extraction kit (Quiagen). A set of primers was designed and their specificity was assessed. MX PCR detected the causal agent in specimens from which T richophyton rubrum and T . interdigitale grew in culture and also identified a dermatophyte species in an additional 32 specimens that were negative in microscopy and culture. None of the investigated non‐dermatophytic strains was positive. Sensitivity of MX PCR was higher as compared to mycological examination (97% vs. 81.1%). MX PCR for direct detection of dermatophytes from nail samples yielded mixed flora in 32.8% of samples. MX PCR proved sensitive and adequate for the diagnosis of dermatophytic onychomycosis. It is much adapted to cases where culture is negative or contaminated by overgrowing moulds, which makes the identification of the causal agent problematic.