Molecular Diagnostic Techniques for Onychomycosis: Validity and Potential Application.
BACKGROUND
Diagnosis of onychomycosis requires positive findings by direct microscopy and fungal culture. Fungal culture is slow and difficult, with low yields. We compared two dermatophyte identification methods, one using a real-time polymerase chain reaction (PCR) method, and the other using fungal culture to validate the molecular method.


METHODS
Nail specimens were collected from 149 patients with distal and lateral subungual onychomycosis who were positive for fungal elements by direct microscopy using potassium hydroxide. Each specimen was subjected to the modified real-time PCR assay of Miyajima et al. and fungal culture.


RESULTS
Of 149 specimens, 142 (95.3%) were positive for Trichophyton rubrum or Trichophyton mentagrophytes including Trichophyton interdigitale by PCR, while only 104 (69.8%) were positive by fungal culture performed simultaneously. No specimen was negative by PCR, but positive by culture. All specimens positive for T. rubrum or T. mentagrophytes by culture were also positive by PCR, showing complete concordance for Trichophyton species. The culture of 17 specimens yielded fungi other than T. rubrum or T. mentagrophytes, whereas PCR identified T. rubrum in 11 of these specimens. Among 28 culture-negative specimens, 23 showed T. rubrum and four showed T. mentagrophytes by PCR. PCR allowed more rapid identification of causative fungi (≤2 days vs. ≤28 days).


CONCLUSIONS
Real-time PCR achieved a higher dermatophyte identification rate and showed complete concordance with conventional culture for two Trichophyton species. Specimens never yielded both T. mentagrophytes and T. rubrum simultaneously, suggesting that mixed infection is uncommon.
Department of Dermatology, Teikyo University School of Medicine, 11-1. Kaga-2, Itabashi, Tokyo, 173-8605, Japan. watanabe@med.teikyo-u.ac.jp.
Shinichi
S
Watanabe
Basic Research Laboratories, Hisamitsu Pharmaceutical Co., Inc., Tsukuba, Ibaraki, Japan.
Kazuya
K
Ishida
D018929:Cell Culture Techniques; D004271:DNA, Fungal / Q000302:isolation & purification*; D006801:Humans; D008853:Microscopy; D025202:Molecular Diagnostic Techniques; D009262:Nails / Q000000981:diagnostic imaging / Q000382:microbiology; D014009:Onychomycosis / Q000175:diagnosis* / Q000382:microbiology*; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013048:Specimen Handling; D013997:Time Factors; D014249:Trichophyton / Q000235:genetics / Q000302:isolation & purification*
D004271:DNA, Fungal