• Users Online: 230
  • Print this page
  • Email this page
  • Email this page
  • Email this page
  • Email this page

Table of Contents
Year : 2019  |  Volume : 37  |  Issue : 2  |  Page : 63-66

Methods for diagnosing onychomycosis: Acomparative study of 459cases

1 Department of Dermatology, Mackay Memorial Hospital; Department of Medicine, Mackay Medical College, New Taipei City; Department of Cosmetic Science and Management, Mackay Medicine, Nursing and Management College, Taipei, Taiwan
2 Deapartment of Dermatology, Taipei Chang Gung Memorial Hospital, Taipei; Department of Medicine, Chang Gung University College of Medicine, Taoyuan, Taiwan
3 Medical Research, Mackay Memorial Hospital, Taipei, Taiwan
4 Department of Dermatology, Mackay Memorial Hospital; Department of Medicine, Mackay Medical College, New Taipei City, Taipei, Taiwan

Date of Submission27-Mar-2018
Date of Acceptance15-Aug-2018
Date of Web Publication23-May-2019

Correspondence Address:
Dr. Yu-Hung Wu
Department of Dermatology, Mackay Memorial Hospital, 92, Sec 2, Zhongshan North Road, Taipei 10449
Login to access the Email id

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ds.ds_6_18

Rights and Permissions

Background: Onychomycosis is a common infectious nail problem but shares similar clinical presentation with some other noninfectious disorders, such as psoriasis and lichen planus. Acorrect diagnosis is essential to proper management. There are three common tests used in the outpatient clinic, including direct potassium hydroxide(KOH) examination, nail plate histology study, and fungal culture. Objective: This study aimed to compare the accuracy and feasibility among these three tests in the diagnosis of onychomycosis. Materials and Methods: We retrospectively reviewed the patients diagnosed with onychomycosis and had positive result of any of the three tests from June 2005 to June 2015. The age, gender, and results of the diagnostic methods were collected and analyzed. Results: Atotal of 459patients were enrolled. The positive rates were significantly different between nail plate histology study(93.0%, 427/459), KOH examination(67.3%, 309/459), and fungal culture(42.1%, 193/459)(P<0.05). Conclusions: Nail plate histology study has the highest positive rate in the diagnosis of onychomycosis than KOH examination and fungal culture. However, KOH examination is most inexpensive and is the fast way to obtain the result with moderate reliability. Acombination of two or three diagnostic tests could provide useful information to the clinician to manage onychomycosis appropriately.

Keywords: Fungal culture, hydroxide examination, nail plate histological examination, onychomycosis

How to cite this article:
Lin YC, Sun PL, Hsiao PF, Sun FJ, Wu YH. Methods for diagnosing onychomycosis: Acomparative study of 459cases. Dermatol Sin 2019;37:63-6

How to cite this URL:
Lin YC, Sun PL, Hsiao PF, Sun FJ, Wu YH. Methods for diagnosing onychomycosis: Acomparative study of 459cases. Dermatol Sin [serial online] 2019 [cited 2023 May 29];37:63-6. Available from: https://www.dermsinica.org/text.asp?2019/37/2/63/258944

  Introduction Top

Onychomycosis is characterized by subungual hyperkeratosis and discoloration of the nail plates. Several other nail disorders, such as psoriasis, lichen planus, twenty-nail dystrophy, might show similar clinical features as onychomycosis. To avoid the inappropriate use of antifungal therapy, the diagnosis of onychomycosis should be confirmed before the initiation of therapeutic agents. There are three commonly used tests in the outpatient clinic, the potassium hydroxide(KOH) direct microscopic observation technique, the pathological examination of nail plate tissue, and fungal culture of nail plate fragment or debris.[1],[2] The KOH is easy to perform and can obtain the result in minutes. The nail plate histology test takes days to get the result. The culture usually required weeks to identify the fungus species. This study aimed to understand the positive rate among these three methods.

  Materials and Methods Top

We retrospectively reviewed the results of direct KOH examination, nail plate histology study, and nail fungal culture among patients who have been diagnosed as onychomycosis between June 2005 and June 2015. The study was approved by the Institutional Review Board(16MMHIS051). The patients with the diagnosis of onychomycosis were retrieved by ICD-9 code(110.1). The records of direct KOH examination, nail plate histology study, and nail fungal culture were collected in the medical records. The patients received all three tests and had at least one positive result of the three diagnostic tests were included in this study. The age, gender, and result of the diagnostic methods were collected and analyzed.

Potassium hydroxide examination

The surface of the nail plate or subungual keratin debris was scraped by using a number 15 scalpel blade. The test materials were placed on the microscope slides. Asmall amount of 20% KOH solution was placed on the periphery of the coverslip. The slide was gently heated for 10 s and then rested for 3min. The slides were examined and reported by experienced residents.

Pathological examination

Nail specimens for histology examination were obtained with the use of standard nail clippers in the following manner: The distal free edge of the nail plate, along with any attached subungual debris, was clipped just distal to its attachment to the nail bed. The clipping was then placed in 10% formalin solution and processed in routine fashion. After processing the specimens, they were stained with periodic acid–Schiff(PAS) stain and evaluated microscopically by a dermatopathologist(Wu) for the presence of any fungal elements. The location and amount were record. The presence of hyphae in the nail plate and/or subungual hyperkeratosis or presence of large amounts of spores in the subungual area was regarded as positive result. Very small sporadic spores indicated contamination or colonization will be reported as negative result.

Fungal culture

The keratin and debris of subungual curettage were cultured on Sabouraud dextrose agar with chloramphenicol and cycloheximide. Cultures were maintained for 4weeks and checked periodically for growth by a mycologist(Sun). Cultures were considered positive if a dermatophyte grew. Culture yields of nondermatophytes or yeasts were considered positive if the same organisms grew on a second subculture from the first inoculated tube or the mold feature consistent with organisms appearing in the histology sample.


We use Cochran's Q statistic test and McNemar test with Bonferroni posthoc correction to assess the statistical significance difference between three tests and paired tests.

  Results Top

A total of 459patients were enrolled in this study, including 194males(42.3%) and 265females(57.7%). The mean age was 45.9±17.3years(ranged from 3 to 91years old). Most of the examined sites are toes(402/459, 87.6%), followed by fingers(57/459, 12.4%).

The percentage of performed tests

A total of 127(27.7%) patients had positive results in three tests, 216(47.0%) patients had two positive tests, and 116(25.3%) patients had only one positive test. The result was summarized in [Table1].
Table 1: Summary of tests results

Click here to view

The positive rate in three different tests

The positive rates were 93.0%(427/459) in nail plate histology study, 67.3%(309/459) in KOH examination, and 42.1%(193/459) in fungal culture. The Cochran's Q statistic test showed a significant difference between these three tests(Cochran's Q=48.763, P <0.05). The McNemar test also showed a significant difference between each test(P=0.017).

Fungal species identified in the fungal culture

A total of 193cases had positive fungal culture results. The most common one was dermatophytes(156/193, 80.8%), followed by mold(19/193, 9.8%), and yeasts(18/193, 9.3%). The detailed species and their percentage were listed in [Table2].
Table 2: Fungal organisms identified among 193 positive culture results

Click here to view

The positivity in different age group

Eight child patients(equal or<12years old) of onychomycosis(8/459, 1.7%) were identified, including five toenails and three fingernails infection. In patients older than 12years old, there are more toenail infection(397/451, 88%) than fingernail infection(54/459, 12%)(P<0.05). However, when we compared the different species in these age groups, there was no significant difference between dermatophytes, yeasts, and nondermatophyte group(P>0.05).

  Discussion Top

This study demonstrated the highest diagnostic rate of histologic evaluation. The PAS stain has a great contrast to identify the fungal element easily[Figure1]. The procedure of nail clipping is easy to perform in a busy clinic. The disadvantages in this technique include taking days to obtain the results, experienced dermatopathologists or pathologists are required to avoid false-positive or false-negative results, could not identify the species and fungi, and highest cost compared to other two methods [Table3].[3],[4],[5] The health insurance in Taiwan reimburses New Taiwan dollars(NT$) 2820 for histologic evaluation(including the clipping technique, specimen processing, and stain), NT$90 for KOH preparation, and NT$330 for fungal culture.[3]
Figure1: The nail plate histology examination may show some different fungal species characteristics in onychomycosis.(a) Fusarium with large chlamydospores(arrow).(b) Aspergillus with acute angle branching hyphae(arrow).(c) Candida Species with budding spores(arrow). (d) Fungal elements in Scedosporium apiosperium not diagnostic for species.(Periodic acid–Schiff stain, a-d, ×400)

Click here to view
Table 3: The clinical usefulness and performance of each test

Click here to view

Histology study requires that the nails be fixed, dehydrated, embedded in paraffin, and sectioned before stain, it takes at least several days to gain the result. Apitfall of this technique is the specimen can only be clipped from the distal part of nail plate and may yield false-negative result if the fungi were located in the proximal nail plate. For suspicious proximal subungual onychomycosis, the healthy nail plate needed to be pared away with a number 15 blade or a sharp curette and then collected materials from the subungual area or nail bed close to the lunula. Alternatively, a superficial punch biopsy of the proximal nail plate can be taken from the proximal onycholytic border without anesthesia.[6] However, it takes more times to perform the procedure.

In the practice, the KOH examination is the rapid and inexpensive ways to diagnosis onychomycosis.[3] The method only provides the presence of fungal elements in the scrapped skin or not and cannot determine whether the fungi were true pathogens or contamination.[5] The KOH examination has been reported to have a false-negative percentage between 5% and 15%, depending on the skill and experience of the performer.[2],[4]

Fungal culture is a highly specific technique, but it has a very high false negative rate, which impedes its sensitivity.[5] One relevant factor was that most specimens were collected from the distal portion, in which the fungus may be old and difficult to grow in culture medium.[7] In addition, fungi requires several weeks to grow in the media. Contaminated yeasts, molds, or nondermatophytes may grow faster than dermatophytes, and repeated culture may be required to confirm the pathogens.[5]

Mold infection of nails has few specific clinical features and is often difficult to cure. For the reason, mycological and histological examinations should be performed on any patients with unusual presentation or therapy resistance.[1] Histological examination was helpful to demonstrate the location of fungal invasion, the presence of hyphae or spores, and the fungi were pigmented or nonpigmented.[8] Some specific morphological changes can also be identified in the help of mycological confirmation. For example, large chlamydospores can be observed in Fusarium infection[Figure1]a. Dichotomous branching hyphae with at acute angle of about 45° can be present in Aspergillus infection[Figure1]b.[9] The budding spores is a feature in Candida species[Figure1]c. However, the findings were usually not specific enough to identify the species[Figure1]d.

Candida species were reported to be the cause of 79% fingernail infections in women, but only 21% of fingernail infection in men.[8] In our study, all nine Candida infections are present in the fingernails and 6(67%) were in women. Our findings also support that Candida onychomycosis is more common in fingers of females.[10]

However, the gender difference was not present between yeasts, dermatophytes, and mold group(P>0.05).

Onychomycosis due to molds has been reported to occur on dystrophic nails, especially in older patients.[10] However, the infectious rate of mold did not show significant difference with dermatophyte or yeast in the average age. In the other way, there is a higher rate of fingernail infection(3/8, 37.5%) in the children 12years old or younger compared to the older persons(12.4%)(P<0.05).

The limitation in his study included only patient with a positive diagnostic result and did not enroll false-negative patients. Therefore, we cannot calculate the true sensitivity, specificity, positive and negative predictive value, and positive and negative likelihood ratio. The new diagnostic technique, such as using calcofluor-white stain in KOH examination and molecular diagnosis of species by using polymerase chain reaction, was not included in our study. The presence of fungal element in any of these methods can be saprophyte or contamination. Perform different tests or repeated the test may be required to confirm the infection.

  Conclusions Top

Nail plate histology study has the highest positive rate in the diagnosis of onychomycosis but is time-consuming. The KOH examination is the fastest and cheapest method but lower sensitivity. The fungal culture can show the species of fungi and helps the treatment adjustment if there is a poor response. However, it took the longest time to obtain the result and has the lowest positive rate. Acombination of two or three diagnostic tests yields the best diagnostic rate in the practice.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

AmeenM, LearJT, MadanV, Mohd MustapaMF, RichardsonM. British Association of Dermatologists' guidelines for the management of onychomycosis 2014. Br J Dermatol 2014;171:937-58.  Back to cited text no. 1
WeinbergJM, KoestenblattEK, TutroneWD, TishlerHR, NajarianL. Comparison of diagnostic methods in the evaluation of onychomycosis. JAm Acad Dermatol 2003;49:193-7.  Back to cited text no. 2
AmirI, FoeringKP, LeeJB. Revisiting office-based direct microscopy for the diagnosis of onychomycosis. JAm Acad Dermatol 2015;72:909-10.  Back to cited text no. 3
BonifazA, Rios-YuilJM, ArenasR, AraizaJ, Fernández R, Mercadillo-Pérez P, etal. Comparison of direct microscopy, culture and calcofluor white for the diagnosis of onychomycosis. Rev Iberoam Micol 2013;30:109-11.  Back to cited text no. 4
GuptaAK, SimpsonFC. Diagnosing onychomycosis. Clin Dermatol 2013;31:540-3.  Back to cited text no. 5
EismanS, SinclairR. Fungal nail infection: Diagnosis and management. BMJ 2014;348:g1800.  Back to cited text no. 6
MidgleyG, MooreMK, CookJC, PhanQG. Mycology of nail disorders. JAm Acad Dermatol 1994;31:S68-74.  Back to cited text no. 7
GreerDL. Evolving role of nondermatophytes in onychomycosis. Int J Dermatol 1995;34:521-4.  Back to cited text no. 8
KradinRL, MarkEJ. The pathology of pulmonary disorders due to Aspergillus spp. Arch Pathol Lab Med 2008;132:606-14.  Back to cited text no. 9
LimJT, ChuaHC, GohCL. Dermatophyte and non-dermatophyte onychomycosis in Singapore. Australas J Dermatol 1992;33:159-63.  Back to cited text no. 10



  [Table1], [Table2], [Table3]

This article has been cited by
1 Comparison of Fungal Fluorescent Staining and ITS rDNA PCR-based Sequencing with Conventional Methods for Diagnosis of Onychomycosis
Amal F. Makled, Mabrouk M. Ghonaim, Sahar A.M. Ali, Sally Mohammed ElHefnawy, Mona Salah Sabal, Asmaa Mohammed Elbrolosy
Journal of Pure and Applied Microbiology. 2022;
[Pubmed] | [DOI]
2 A descriptive study of onychoscopic features in various subtypes of onychomycosis
A. Devi Sangeetha,K. Gopalakrishnan,R. Ramachandran,Murali Narasimhan,Balaji Ramraj
Medical Journal Armed Forces India. 2021;
[Pubmed] | [DOI]


    Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
    Access Statistics
    Email Alert *
    Add to My List *
* Registration required (free)  

  In this article
Materials and Me...
Article Figures
Article Tables

 Article Access Statistics
    PDF Downloaded632    
    Comments [Add]    
    Cited by others 2    

Recommend this journal