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The Role of Trypan Blue as a Conventional and Fluorescent Dye for the Diagnosis of Superficial Mycoses by Potassium Hydroxide (KOH) Testing

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Abstract

Direct microscopic examination of samples using potassium hydroxide (KOH) is a fast, simple, and inexpensive method to confirm clinical suspicion of superficial mycosis. However, the lack of color contrast in this test makes it difficult to separate any fungal structures from artifacts. The sensitivity of the KOH mount technique may be enhanced using both fluorochromes and conventional stains that highlight the fungal structures when observed under fluorescence microscopy and bright-field, respectively. Here we study the possibility of using Trypan Blue (TB), an azo dye which is often used as a live/dead marker, in the diagnosis of superficial mycoses by KOH testing. TB at 0.01% displayed a fluorescent staining pattern similar to that of Calcofluor White (CFW), the conventional cell wall fluorophore. Furthermore, by adjusting the TB concentration to 0.1–0.3%, in addition to maintaining the fluorescent staining pattern, the fungal elements were stained in blue under bright-field microscopy. Thus, we demonstrate for the first time that TB has the unique property as a fungal stain that can be used in both bright-field and fluorescence microscopy for diagnosis of superficial mycoses by direct microscopic examination.

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References

  1. Sharma B, Nonzom S. Superficial mycoses, a matter of concern: global and Indian scenario-an updated analysis. Mycoses. 2021. https://doi.org/10.1111/myc.13264.

    Article  PubMed  PubMed Central  Google Scholar 

  2. Pihet M, Le Govic Y. Reappraisal of conventional diagnosis for dermatophytes. Mycopathologia. 2017. https://doi.org/10.1007/s11046-016-0071-y.

    Article  PubMed  Google Scholar 

  3. Ghannoum M, Mukherjee P, Isham N, Markinson B, Rosso JD, Leal L. Examining the importance of laboratory and diagnostic testing when treating and diagnosing onychomycosis. Int J Dermatol. 2018. https://doi.org/10.1111/ijd.13690.

    Article  PubMed  Google Scholar 

  4. Wilkison BD, Sperling LC, Spillane AP, Meyerle JH. How to teach the potassium hydroxide preparation: a disappearing clinical art form. Cutis. 2015;96:109–12.

    PubMed  Google Scholar 

  5. Lim SS, Ohn J, Mun JH. Diagnosis of onychomycosis: from conventional techniques and dermoscopy to artificial intelligence. Front Med (Lausanne). 2021. https://doi.org/10.3389/fmed.2021.637216.

    Article  PubMed  Google Scholar 

  6. Ikeda I, Nishimoto K, Sasamoto K, Nagira K, Ono T, Ihn H. Alkaline Trypan Blue as a stain for superficial fungi. Br J Dermatol. 2008. https://doi.org/10.1111/j.1365-2133.2008.08523.x.

    Article  PubMed  Google Scholar 

  7. Liesche J, Marek M, Günther-Pomorski T. Cell wall staining with Trypan blue enables quantitative analysis of morphological changes in yeast cells. Front Microbiol. 2015. https://doi.org/10.3389/fmicb.2015.00107.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Yue X, Wang A, Wang H, Li Q, Yue X. Evaluation of a new fluorescent reagent, fluorescent brightener 85, for the diagnosis of suspected onychomycosis compared with potassium hydroxide. Mycoses. 2018. https://doi.org/10.1111/myc.12733.

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

I.T.J.F. received a Master’s scholarship from CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior) and B.A.M. received an undergraduate research scholarship from FAPEMIG (Fundação de Amparo à Pesquisa do Estado de Minas Gerais)

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No funding sources. The authors have no funding to disclose.

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Authors and Affiliations

  1. Microbiology Research Laboratory, Institute of Biological and Natural Sciences, Federal University of Triângulo Mineiro, Uberaba, Minas Gerais, Brazil

    Izabella Thays Jacob Felix, Beatriz Arfelli Martins, Celso Tadeu Barbosa dos Santos & Anderson Assunção Andrade

  2. Dermatology Unit, Institute of Health Sciences, Federal University of Triângulo Mineiro, Uberaba, Minas Gerais, Brazil

    Romes José Tristão & Meire Soares de Ataíde

  3. Physiology Laboratory, Institute of Biological and Natural Sciences, Federal University of Triângulo Mineiro, Uberaba, Minas Gerais, Brazil

    Aldo Rogelis Aquiles Rodrigues

  4. Departamento de Microbiologia, Imunologia e Parasitologia, Instituto de Ciências Biológicas e Naturais, Universidade Federal do Triângulo Mineiro (UFTM), Praça Manoel Terra, 330 – Abadia, Uberaba, MG, 38025-015, Brazil

    Anderson Assunção Andrade

Authors
  1. Izabella Thays Jacob Felix
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  2. Beatriz Arfelli Martins
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  3. Celso Tadeu Barbosa dos Santos
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  4. Romes José Tristão
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  5. Aldo Rogelis Aquiles Rodrigues
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  6. Meire Soares de Ataíde
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  7. Anderson Assunção Andrade
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Contributions

ITJF, BAM, and CTBS collected the data. RJT and MSA provided and cared for study patients. ITJF, CTBS, ARAR and AAA analyzed the data. AAA conceived and designed the study, and led the writing, with contributions from ARAR All the authors have read and approved the final manuscript.

Corresponding author

Correspondence to Anderson Assunção Andrade.

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Conflict of interest

The authors have no relevant financial or non-financial interests to disclose.

Ethical Approval

This study was approved by the Ethics Committee of UFTM (protocol 2.623.027).

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Informed consent was obtained from all individual participants included in the study.

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Handling Editor: Jianjun Qiao.

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Felix, I.T.J., Martins, B.A., dos Santos, C.T.B. et al. The Role of Trypan Blue as a Conventional and Fluorescent Dye for the Diagnosis of Superficial Mycoses by Potassium Hydroxide (KOH) Testing. Mycopathologia 188, 815–819 (2023). https://doi.org/10.1007/s11046-023-00771-9

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  • DOI: https://doi.org/10.1007/s11046-023-00771-9

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