Ann Dermatol.  2011 Aug;23(3):304-312. 10.5021/ad.2011.23.3.304.

Identification of Dermatophytes Using Multiplex Polymerase Chain Reaction

• Kim JY
• Choe YB
• Ahn KJ
• Lee YW

• KMID: 2171921
• DOI: http://doi.org/10.5021/ad.2011.23.3.304

Abstract

BACKGROUND
Multiplex polymerase chain reaction (PCR) allows more than two target DNA molecules to be amplified with more than two primers. This method is also useful for detecting various other organisms simultaneously within a single test tube, and the scope of its use has been expanding widely in the field of clinical microbiology in recent years.
OBJECTIVE
To assess the value of multiplex PCR in identification of dermatophytes.
METHODS
Using three specially-designed primers which contained the ITS1-2, 18S rRNA, and 28S rRNA regions, three cycles of PCR were performed on 11 standard strains and scales were collected from 73 patients with fungal infection.
RESULTS
The 11 standard strains were successfully identified with analysis of band patterns of ITS1-2, 18S rRNA, and 28S rRNA, obtained from PCR. Based on this information, the causative organisms in 73 patients with fungal infection were revealed to be T. rubrum in 69 cases, T. menta in 1 case, T. tonsurans in 2 cases, and M. gypseum in one case.
CONCLUSION
With three cycles of PCR using three sets of primers, 11 standard strains and the clinical strains from 73 patients with fungal infection were successfully identified.

Keyword

Dermatophyte; Multiplex PCR

MeSH Terms

Arthrodermataceae
DNA
Humans
Multiplex Polymerase Chain Reaction
Polymerase Chain Reaction
Weights and Measures
DNA

Figure

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  Fig. 1 PCR amplification from dermatophyte standard strains with (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region. Lanes: M, molecular marker; 1. Epidermophyton floccosum (CBS 358.93); 2. Microsporum canis (IFM 45829); 3. Microsporum audouinii (IFM 5294 [CBS 319.31]); 4. Trichophyton violaceum (CBS 319.31); 5. Trichophyton rubrum (KCTC 6375(ATCC 28188)); 6. Trichophyton mentagrophytes var. mentagrophytes (CBS 126.34); 7. Trichophyton tonsurans (CBS 483.76); 8. Trichophyton mentagrophytes var. interdigitale (IFM 53931); 9. Trichophyton verrucosum (CBS 134.66); 10. Microsporum gypseum (IFM 5292); 11. Microsporum fulvum (IFM 5318); 12. Control (dH2O). PCR: polymerase chain reaction.

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  Fig. 2 PCR amplification of (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region from onychomycosis. Lanes: M, molecular marker; 1~24. Trichophyton rubrum. PCR: polymerase chain reaction.

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  Fig. 3 PCR amplification of (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region from tinea pedis. Lanes: M, molecular marker; 3. Trichophyton mentagrophytes var. mentagrophytes, 7. Trichophyton tonsurans. The other lanes were Trichophyton rubrum. PCR: polymerase chain reaction.

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  Fig. 4 PCR amplification of (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region from tinea capitis. Lanes: M, molecular marker; 17~18. Trichophyton rubrum, 19. Trichophyton tonsurans. PCR: polymerase chain reaction.

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  Fig. 5 PCR amplification of (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region from tinea corporis. Lanes: M, molecular marker; 7. Microsporum gypseum. The other lanes were Trichophyton rubrum. PCR: polymerase chain reaction.

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  Fig. 6 PCR amplification of (A) the ITS1-2 region, (B) the 18S ribosomal RNA region and (C) the 28S ribosomal RNA region from tinea cruris. Lanes: M, molecular marker; 1~11. Trichophyton rubrum. PCR: polymerase chain reaction.

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