Development and evaluation of a pan‐dermatophyte PCR with species‐level identification using sloppy molecular beacon probes

Summary

Background

Conventional laboratory diagnosis of dermatophyte infection is cumbersome and time‐consuming.

Objectives

We aimed to establish a simple, robust, and rapid molecular diagnostic assay for detection of dermatophytes and optionally non‐dermatophytes in clinical specimens.

Methods

We developed a two‐tube pan‐dermatophyte PCR assay using six sloppy molecular beacon (SMB) probes. The first PCR uses dermatophyte specific primers and enables detection and identification of most dermatophyte species. The second PCR with pan‐fungal primers allows further differentiation of T. interdigitale and T. mentagrophytes/T. quinckeanum, T. violaceum and T. soudanense, and T. tonsurans and T. equinum, and detection of non‐dermatophytes. The test was evaluated with 306 clinical specimens by comparing it to the results of microscopy and culture.

Results

In melting curve analyses, species‐specific T_m signatures of the SMBs were defined. Thus, our new PCR enabled detection and species‐level identification of at least 19 dermatophyte species. Sensitivity and specificity of PCR for detection of dermatophytes in clinical samples were estimated to be 96.9% and 90.4%, for culture 46.7% and 98.7%, and for microscopy 91.4% and 84.0%, respectively. The detection of non‐dermatophytes by PCR and culture did not correlate.

Conclusions

The new assay showed excellent performance characteristics for the detection of dermatophytes and is significantly faster than culturing techniques what makes it very promising for routine diagnostics of dermatophytosis. We noticed that the detection of non‐dermatophytes in our assay currently has no benefit.

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