Five-Hour Diagnosis of Dermatophyte Nail Infections

24 A rapid two step DNA extraction method and a multiplex PCR for the detection of 25 dermatophytes in general and Trichophyton (T.) rubrum specifically were developed and 26 evaluated on DNA extracted from pure cultures and from clinically diseased nails. DNA 27 from the following dermatophytes were used: Epidermophyton floccosum, Microsporum 28 (M.) audouinii, M. canis, M. gypseum, M. nanum, T. mentagrophytes, T. rubrum, T. 29 schoenleinii, T. soudanense, T. terrestre, T. tonsurans, T. verrucosum and T. violaceum. 30 Human DNA and DNA from the following non-dermatophyte fungi were included as 31 controls: Alternaria, Aspergillus niger, Candida (C.) albicans, C. glabrata, C. krusei, 32 Malassezia furfur, Saccharomyces cerevisiae, and Scopulariopsis brevicaulis. A total of 33 118 nail samples received for routine microscopy and culture for dermatophytes were 34 subsequently tested by the two PCRs separately and in a multiplex format. Using DNA 35 extracted from pure cultures the pan-dermatophyte PCR, the T. rubrum specific PCR 36 sequentially and in a multiplex format correctly detected all dermatophytes and 37 additionally correctly identified T. rubrum. Comparison of the traditional diagnostic 38 evaluation (microscopy and culture) of nail samples with PCR on DNA directly extracted 39 from the nails showed excellent agreement between PCR and microscopy, but the 40 number of samples with dermatophyte species identification was increased considerably 41 from 22.9% to 41.5%, mainly due to the identification of T. rubrum by PCR in 42 AC CE PT ED on A uust 5, 2017 by gest ht://jcm .sm .rg/ D ow nladed fom microscopy positive but culture negative samples. In conclusion, this 5-hour diagnostic 43 test was shown to increase not only speed but also sensitivity of investigation for nail 44 dermatophytosis. 45