PCR-reverse blot hybridization assay for species identification of dermatophytes

Hyunjung Kim, Hyunwoo Jin, Sunghyun Kim, Hye Young Wang, Yeonim Choi, Hyeeun Bang, Je Seop Park, Jang Ho Lee, Young Ho Won, Kyu Joong Ahn, Young Kwon Kim, Hyeyoung Lee

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Background: Dermatophytes (Trichophyton, Microsporum, and Epidermophyton) cause cutaneous mycoses called dermatophytosis. Forproper anti-dermatophytosis therapy, accurate and early diagnosis of dermatophytes is important. Laboratory diagnosis of dermatophytosis for dermatophytes still relies on microscopic and macroscopic examination of in vitro cultures and some physiological tests. These methods (conventional methods) are time-consuming (2-4 weeks) and yet, still have low sensitivity and specificity. Recently, in order to overcome such limitations of conventional methods, molecular-based methods have been developed to identify dermatophytes. The polymerase chain reaction-reverse blot hybridization assay (PCR-REBA) allows sensitive and specific identification of dermatophytes species. Objective: This study was aimed to develop a new PCR-REBA with higher sensitivity using less amount of probe concentration, so the assay can be more practical in clinical settings. Methods: For this, PCR primers and species-specific oligonucleotide probes were designed within the internal transcribed sequences 1 region between 5.8S and 18S rRNA. The species-specific probes designed in this study was to identify 6 species (T. rubrum, T. mentagrophytes, T. tonsurans, M. canis, M. gypseum, and E. floccosum) comprised 99% of dermatophytes isolatedin Korea. Results: The detection efficiency of the PCR-REBA was compared with the microscopic method, and the results showed that the sensitivity of the PCR-REBA developed in this study is 100 times higher than previously developed one. Subsequently, the results of PCR-REBA were evaluated using clinical isolates. DNAs from a total of 68 clinical isolates were analyzed by PCR-REBA, and the inconsistent results between PCR-REBA and conventional microscopic identification results were confirmed by sequence analysis. Conclusion: In brief, the results showed that results of sequence analysis were identical with PCR-REBA implying newly developed PCR-REBA is very useful method for accurate and rapid identification of dermatophytes and would provide higher simplicity, specificity, sensitivity than conventional method.

Original languageEnglish
Pages (from-to)86-98
Number of pages13
JournalKorean Journal of Medical Mycology
Volume16
Issue number3
DOIs
Publication statusPublished - 2011 Jan 1

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Arthrodermataceae
Polymerase Chain Reaction
Tinea
Sequence Analysis
Epidermophyton
Microsporum
Sensitivity and Specificity
Trichophyton
Oligonucleotide Probes
Mycoses
Clinical Laboratory Techniques
Korea
Early Diagnosis
Skin

All Science Journal Classification (ASJC) codes

  • Infectious Diseases

Cite this

Kim, Hyunjung ; Jin, Hyunwoo ; Kim, Sunghyun ; Wang, Hye Young ; Choi, Yeonim ; Bang, Hyeeun ; Park, Je Seop ; Lee, Jang Ho ; Won, Young Ho ; Ahn, Kyu Joong ; Kim, Young Kwon ; Lee, Hyeyoung. / PCR-reverse blot hybridization assay for species identification of dermatophytes. In: Korean Journal of Medical Mycology. 2011 ; Vol. 16, No. 3. pp. 86-98.
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abstract = "Background: Dermatophytes (Trichophyton, Microsporum, and Epidermophyton) cause cutaneous mycoses called dermatophytosis. Forproper anti-dermatophytosis therapy, accurate and early diagnosis of dermatophytes is important. Laboratory diagnosis of dermatophytosis for dermatophytes still relies on microscopic and macroscopic examination of in vitro cultures and some physiological tests. These methods (conventional methods) are time-consuming (2-4 weeks) and yet, still have low sensitivity and specificity. Recently, in order to overcome such limitations of conventional methods, molecular-based methods have been developed to identify dermatophytes. The polymerase chain reaction-reverse blot hybridization assay (PCR-REBA) allows sensitive and specific identification of dermatophytes species. Objective: This study was aimed to develop a new PCR-REBA with higher sensitivity using less amount of probe concentration, so the assay can be more practical in clinical settings. Methods: For this, PCR primers and species-specific oligonucleotide probes were designed within the internal transcribed sequences 1 region between 5.8S and 18S rRNA. The species-specific probes designed in this study was to identify 6 species (T. rubrum, T. mentagrophytes, T. tonsurans, M. canis, M. gypseum, and E. floccosum) comprised 99{\%} of dermatophytes isolatedin Korea. Results: The detection efficiency of the PCR-REBA was compared with the microscopic method, and the results showed that the sensitivity of the PCR-REBA developed in this study is 100 times higher than previously developed one. Subsequently, the results of PCR-REBA were evaluated using clinical isolates. DNAs from a total of 68 clinical isolates were analyzed by PCR-REBA, and the inconsistent results between PCR-REBA and conventional microscopic identification results were confirmed by sequence analysis. Conclusion: In brief, the results showed that results of sequence analysis were identical with PCR-REBA implying newly developed PCR-REBA is very useful method for accurate and rapid identification of dermatophytes and would provide higher simplicity, specificity, sensitivity than conventional method.",
author = "Hyunjung Kim and Hyunwoo Jin and Sunghyun Kim and Wang, {Hye Young} and Yeonim Choi and Hyeeun Bang and Park, {Je Seop} and Lee, {Jang Ho} and Won, {Young Ho} and Ahn, {Kyu Joong} and Kim, {Young Kwon} and Hyeyoung Lee",
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Kim, H, Jin, H, Kim, S, Wang, HY, Choi, Y, Bang, H, Park, JS, Lee, JH, Won, YH, Ahn, KJ, Kim, YK & Lee, H 2011, 'PCR-reverse blot hybridization assay for species identification of dermatophytes', Korean Journal of Medical Mycology, vol. 16, no. 3, pp. 86-98. https://doi.org/10.17966/kjmm.2011.16.3.86

PCR-reverse blot hybridization assay for species identification of dermatophytes. / Kim, Hyunjung; Jin, Hyunwoo; Kim, Sunghyun; Wang, Hye Young; Choi, Yeonim; Bang, Hyeeun; Park, Je Seop; Lee, Jang Ho; Won, Young Ho; Ahn, Kyu Joong; Kim, Young Kwon; Lee, Hyeyoung.

In: Korean Journal of Medical Mycology, Vol. 16, No. 3, 01.01.2011, p. 86-98.

Research output: Contribution to journalArticle

TY - JOUR

T1 - PCR-reverse blot hybridization assay for species identification of dermatophytes

AU - Kim, Hyunjung

AU - Jin, Hyunwoo

AU - Kim, Sunghyun

AU - Wang, Hye Young

AU - Choi, Yeonim

AU - Bang, Hyeeun

AU - Park, Je Seop

AU - Lee, Jang Ho

AU - Won, Young Ho

AU - Ahn, Kyu Joong

AU - Kim, Young Kwon

AU - Lee, Hyeyoung

PY - 2011/1/1

Y1 - 2011/1/1

N2 - Background: Dermatophytes (Trichophyton, Microsporum, and Epidermophyton) cause cutaneous mycoses called dermatophytosis. Forproper anti-dermatophytosis therapy, accurate and early diagnosis of dermatophytes is important. Laboratory diagnosis of dermatophytosis for dermatophytes still relies on microscopic and macroscopic examination of in vitro cultures and some physiological tests. These methods (conventional methods) are time-consuming (2-4 weeks) and yet, still have low sensitivity and specificity. Recently, in order to overcome such limitations of conventional methods, molecular-based methods have been developed to identify dermatophytes. The polymerase chain reaction-reverse blot hybridization assay (PCR-REBA) allows sensitive and specific identification of dermatophytes species. Objective: This study was aimed to develop a new PCR-REBA with higher sensitivity using less amount of probe concentration, so the assay can be more practical in clinical settings. Methods: For this, PCR primers and species-specific oligonucleotide probes were designed within the internal transcribed sequences 1 region between 5.8S and 18S rRNA. The species-specific probes designed in this study was to identify 6 species (T. rubrum, T. mentagrophytes, T. tonsurans, M. canis, M. gypseum, and E. floccosum) comprised 99% of dermatophytes isolatedin Korea. Results: The detection efficiency of the PCR-REBA was compared with the microscopic method, and the results showed that the sensitivity of the PCR-REBA developed in this study is 100 times higher than previously developed one. Subsequently, the results of PCR-REBA were evaluated using clinical isolates. DNAs from a total of 68 clinical isolates were analyzed by PCR-REBA, and the inconsistent results between PCR-REBA and conventional microscopic identification results were confirmed by sequence analysis. Conclusion: In brief, the results showed that results of sequence analysis were identical with PCR-REBA implying newly developed PCR-REBA is very useful method for accurate and rapid identification of dermatophytes and would provide higher simplicity, specificity, sensitivity than conventional method.

AB - Background: Dermatophytes (Trichophyton, Microsporum, and Epidermophyton) cause cutaneous mycoses called dermatophytosis. Forproper anti-dermatophytosis therapy, accurate and early diagnosis of dermatophytes is important. Laboratory diagnosis of dermatophytosis for dermatophytes still relies on microscopic and macroscopic examination of in vitro cultures and some physiological tests. These methods (conventional methods) are time-consuming (2-4 weeks) and yet, still have low sensitivity and specificity. Recently, in order to overcome such limitations of conventional methods, molecular-based methods have been developed to identify dermatophytes. The polymerase chain reaction-reverse blot hybridization assay (PCR-REBA) allows sensitive and specific identification of dermatophytes species. Objective: This study was aimed to develop a new PCR-REBA with higher sensitivity using less amount of probe concentration, so the assay can be more practical in clinical settings. Methods: For this, PCR primers and species-specific oligonucleotide probes were designed within the internal transcribed sequences 1 region between 5.8S and 18S rRNA. The species-specific probes designed in this study was to identify 6 species (T. rubrum, T. mentagrophytes, T. tonsurans, M. canis, M. gypseum, and E. floccosum) comprised 99% of dermatophytes isolatedin Korea. Results: The detection efficiency of the PCR-REBA was compared with the microscopic method, and the results showed that the sensitivity of the PCR-REBA developed in this study is 100 times higher than previously developed one. Subsequently, the results of PCR-REBA were evaluated using clinical isolates. DNAs from a total of 68 clinical isolates were analyzed by PCR-REBA, and the inconsistent results between PCR-REBA and conventional microscopic identification results were confirmed by sequence analysis. Conclusion: In brief, the results showed that results of sequence analysis were identical with PCR-REBA implying newly developed PCR-REBA is very useful method for accurate and rapid identification of dermatophytes and would provide higher simplicity, specificity, sensitivity than conventional method.

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