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Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of The Topoisomerase II.
Cell J. 2020 Apr; 22(1):85-91.CJ

Abstract

Objective

Precise identification of dermatophyte species significantly improves treatment and controls measures of dermatophytosis in human and animals. This study was designed to evaluate molecular tools effectiveness of the gene sequencing and DNA-based fragment polymorphism analysis for accurate identification and differentiation of closelyrelated dermatophyte species isolated from clinical cases of dermatophytosis and their antifungal susceptibility to the current antifungal agents.

Materials and Methods

In this experimental study, a total of 95 skin samples were inoculated into mycobiotic agar for two weeks at 28˚C. Morphological characteristics of the isolated dermatophytes were evaluated. DNA was extracted from the fungal culture for amplification of topoisomerase II gene fragments and polymerase chain reaction (PCR) products were digested by Hinf I enzyme. Internal transcribed spacer (ITS) rDNA and TEF-1α regions of the all isolates were amplified using the primers of ITS1/4 and EF-DermF/EF-DermR, respectively.

Results

Based on the morphological criteria, 24, 24, 24 and 23 isolates were identified as T. rubrum , T. interdigitale, T. tonsurans and E. floccosum, respectively. PCR-restriction fragment length polymorphism (RFLP) results provided identification pattern of the isolates for T. rubrum (19 isolates), T. tonsurans (28 isolates), T. interdigitale (26 isolates) and E. floccosum (22 isolates). Concatenated dataset results were similar in PCR-RFLP, except six T. interdigitale isolates belonging to T. mentagrophytes.

Conclusion

Our results clearly indicated that conventional morphology and PCR-RFLP were not able to precisely identify all dermatophyte species and differentiation of closely related species like T. interdigitale and T. mentagrophytes, while ITS rDNA and TEF-1α gene sequence analyses provided accurate identification of all isolates at the genus and species level.

Authors+Show Affiliations

Department of Mycology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.Department of Mycology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran. Electronic Address: shamsm@modares.ac.ir.Department of Mycology, Pasteur Institute of Iran, Tehran, Iran.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

31606971

Citation

Salehi, Zahra, et al. "Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of the Topoisomerase II." Cell Journal, vol. 22, no. 1, 2020, pp. 85-91.
Salehi Z, Shams-Ghahfarokhi M, Razzaghi-Abyaneh M. Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of The Topoisomerase II. Cell J. 2020;22(1):85-91.
Salehi, Z., Shams-Ghahfarokhi, M., & Razzaghi-Abyaneh, M. (2020). Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of The Topoisomerase II. Cell Journal, 22(1), 85-91. https://doi.org/10.22074/cellj.2020.6372
Salehi Z, Shams-Ghahfarokhi M, Razzaghi-Abyaneh M. Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of the Topoisomerase II. Cell J. 2020;22(1):85-91. PubMed PMID: 31606971.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Internal Transcribed Spacer rDNA and TEF-1α Gene Sequencing of Pathogenic Dermatophyte Species and Differentiation of Closely Related Species Using PCR-RFLP of The Topoisomerase II. AU - Salehi,Zahra, AU - Shams-Ghahfarokhi,Masoomeh, AU - Razzaghi-Abyaneh,Mehdi, Y1 - 2019/09/08/ PY - 2018/09/11/received PY - 2019/12/26/accepted PY - 2020/04/01/pmc-release PY - 2019/10/14/entrez PY - 2019/10/14/pubmed PY - 2019/10/14/medline KW - Dermatophytes KW - Gene Sequencing KW - Polymerase Chain Reaction-Restriction Fragment Length Polymorphism KW - Topoisomerase II SP - 85 EP - 91 JF - Cell journal JO - Cell J VL - 22 IS - 1 N2 - Objective: Precise identification of dermatophyte species significantly improves treatment and controls measures of dermatophytosis in human and animals. This study was designed to evaluate molecular tools effectiveness of the gene sequencing and DNA-based fragment polymorphism analysis for accurate identification and differentiation of closelyrelated dermatophyte species isolated from clinical cases of dermatophytosis and their antifungal susceptibility to the current antifungal agents. Materials and Methods: In this experimental study, a total of 95 skin samples were inoculated into mycobiotic agar for two weeks at 28˚C. Morphological characteristics of the isolated dermatophytes were evaluated. DNA was extracted from the fungal culture for amplification of topoisomerase II gene fragments and polymerase chain reaction (PCR) products were digested by Hinf I enzyme. Internal transcribed spacer (ITS) rDNA and TEF-1α regions of the all isolates were amplified using the primers of ITS1/4 and EF-DermF/EF-DermR, respectively. Results: Based on the morphological criteria, 24, 24, 24 and 23 isolates were identified as T. rubrum , T. interdigitale, T. tonsurans and E. floccosum, respectively. PCR-restriction fragment length polymorphism (RFLP) results provided identification pattern of the isolates for T. rubrum (19 isolates), T. tonsurans (28 isolates), T. interdigitale (26 isolates) and E. floccosum (22 isolates). Concatenated dataset results were similar in PCR-RFLP, except six T. interdigitale isolates belonging to T. mentagrophytes. Conclusion: Our results clearly indicated that conventional morphology and PCR-RFLP were not able to precisely identify all dermatophyte species and differentiation of closely related species like T. interdigitale and T. mentagrophytes, while ITS rDNA and TEF-1α gene sequence analyses provided accurate identification of all isolates at the genus and species level. SN - 2228-5806 UR - https://www.unboundmedicine.com/medline/citation/31606971/Internal_Transcribed_Spacer_rDNA_and_TEF_1α_Gene_Sequencing_of_Pathogenic_Dermatophyte_Species_and_Differentiation_of_Closely_Related_Species_Using_PCR_RFLP_of_The_Topoisomerase_II_ L2 - https://www.ncbi.nlm.nih.gov/pmc/articles/pmid/31606971/ DB - PRIME DP - Unbound Medicine ER -
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