Review

Examining the importance of laboratory and diagnostic

testing when treating and diagnosing onychomycosis
Mahmoud Ghannoum1, PhD, Pranab Mukherjee1, PhD, Nancy Isham1, M(ASCP),
Bryan Markinson2, DPM, James Del Rosso3, DO, and Luis Leal4, DPM

1
Case Western University, Cleveland, OH, Abstract
2
Icahn School of Medicine at Mount Sinai, Onychomycosis is a fungal nail infection caused primarily by dermatophytes. Several other
New York, NY, 3Touro University Nevada,
nail disorders, including psoriasis, can simulate onychomycosis. Accurate diagnosis is
Henderson, NV, and 4PharmaDerm a
division of Fougera Pharmaceuticals Inc.,
therefore vital for the ongoing treatment and management of onychomycosis and to avoid
Princeton, NJ, USA misdiagnosis and treatment delay, which can be both lengthy and costly. Often, a
combination of histologic and laboratory techniques is used to obtain an accurate
Correspondence diagnosis. The potential diagnostic challenges associated with the differential diagnosis of
Mahmoud Ghannoum, PhD
onychomycosis caused by dermatophytes and the most common techniques used to
Case Western University
confirm the diagnosis are discussed.
10900 Euclid Avenue
Cleveland, OH 44106
E-mail: mahmoud.ghannoum@case.edu

Disclosure: Writing and editorial assistance

was provided by ApotheCom, Associates,
LLC, Yardley, PA, and was supported by
Sandoz, a Novartis Division. The authors
were fully responsible for the content,
editorial decisions, and opinions expressed
in the current article. No author received an
honorarium related to the development of
this manuscript. L. Leal is an employee of
Sandoz, a Novartis Division.

doi: 10.1111/ijd.13690

that, in addition to exposure to environmental risk factors, leads

Introduction
Onychomycosis is a fungal infection of the nail caused primarily
by dermatophytes, which include Trichophyton, Epidermophy-
ton, and Microsporum spp.1 Infected patients experience discol-
oration, nail plate thickening, and onycholysis.1 Dermatophytes
are transmitted via contact with floor and ground surfaces and
occasionally through personal contact, including people living in
the same household.2 Toenails are more commonly affected
than fingernails, as they are in direct contact with reservoirs in
which dermatophytes commonly colonize, are often confined to
the moist environment within shoes, and grow more slowly.1
It is well documented that environmental factors are involved
in the establishment of onychomycosis; however, some individu-
als exposed to those same environmental risk factors do not
contract onychomycosis, while others will develop a chronic
infection.3 There is emerging evidence that suggests a genetic
component in the susceptibility of an individual to fungal infec-
tion (e.g., defects in the innate and adaptive immune system)
to chronic onychomycosis.3
This disease is reported to affect approximately 10% of the
general population, with prevalence increasing with age.2 In this
regard, prevalence is approximately 20% in patients over
60 years of age and up to 50% in patients older than 70 years.2
Onychomycosis can be classified into distinct clinical categories
based on the region of the nail unit that is affected. These cate-
gories include distal/lateral onychomycosis, proximal subungual
onychomycosis, and superficial white onychomycosis.4 If left
untreated, onychomycosis will often worsen in severity, leading
to marked dystrophic changes in affected nails. Furthermore,
onychomycosis has been reported to have a significant impact
on patient quality of life due to a variety of physical changes
(e.g., pain, discomfort, difficulty trimming thick nail plates, and
difficulty walking) and psychosocial consequences (e.g., embar-
rassment and avoidance of intimacy).2
As nail dystrophy because of fungal infections represents
50% of clinical manifestations, obtaining an accurate diagnosis 1

ª 2017 The International Society of Dermatology International Journal of Dermatology 2017

2 Review Diagnosing onychomycosis by laboratory testing
is critical when treating onychomycosis in order to avoid misdi-
agnosis and treatment delay.4 This article discusses the impor-
tance of obtaining an accurate diagnosis and reviews the
different laboratory techniques available to confirm the diagno-
sis of onychomycosis caused by dermatophytes.
Differential Diagnosis of Onychomycosis
As mentioned previously, only 50% of nail dystrophy is because
of fungal infection, with several other conditions, including
inflammatory disorders such as psoriasis and lichen planus,
presenting nail changes that clinically mimic onychomycosis.1,5,6
Additionally, the clinical appearances of onychomycosis caused
by different fungal species are often indistinguishable, thus indi-
cating that laboratory testing is required for identification of the
infecting organism.7
History and clinical evaluation are not sufficient to make the
diagnosis of onychomycosis, and empiric treatment of pre-
sumed onychomycosis may lead to lower than expected cure
rates and missed diagnoses. A diagnosis based solely on physi-
cal examination/clinical signs can be incorrect,1,8 and various
laboratory assessments would be useful for confirmation.5
Physicians should be aware that laboratory confirmation,
when used in conjunction with their clinical expertise, can avoid
delays in treatment.9 Traditional diagnostic techniques are
based on microscopy, fungal culture, and histological examina-
tion. New technologies such as molecular assays and mass
spectrometry are expanding the clinical laboratory capabilities to
correctly identify dermatophyte to the species level.
Optimal Specimen Collection Technique
Obtaining clean samples is vital for the diagnosis of onychomy-
cosis, as sample contamination is common. Simple steps to
prevent or minimize sample contamination include the use of a
sterilizing solution such as 70% isopropanol to cleanse the nail
plate and surrounding soft tissue prior to sample collection.1,5 A
sterile nail clipper should be used to clip the nail plate, and a
sterile curette or blade should be used to obtain subungual deb-
ris from underneath the nail plate (i.e., the exposed nail bed
where the infection resides).1
Another critical factor is the collection of enough sample for
microscopic examination and culture,10 as too often, inadequate
nail samples (either in quantity or quality) have led to failure of
fungal diagnosis. The location from which the sample should be
collected varies depending on the category/type of onychomy-
cosis:1 for distal and lateral subungual onychomycosis the sam-
ple should be taken from the most proximal location after
clipping of the distal onycholytic nail plate;11 in proximal subun-
gual onychomycosis, the upper nail plate should be debrided
and a sample containing the underlying nail debris should be
collected; and for superficial white onychomycosis, the speci-
men should be obtained from affected superficial nail plate.1
International Journal of Dermatology 2017
Ghannoum et al.
Samples for the latter two types can be collected with a nail drill
to enhance precision.11 Moreover, the type of sample collected
also depends on the laboratory test being used. Periodic acid–
Schiff (PAS) staining requires nail plate clippings but will not
distinguish fungal species. On the other hand, subungual debris
sent for fungal culture may provide viable fungal elements of
dermatophytes, nondermatophytic molds, and yeasts that can
be identified to genus and species.12,13
Diagnostic Techniques
A diagnosis of onychomycosis is confirmed when fungal hyphae
and fungal viability are demonstrated and the fungal species
identified.14 A variety of laboratory and diagnostic techniques,
described below, are currently used, all of which have benefits
and limitations; thus accurate diagnosis is often achieved using
a combination of these techniques.14
10% Potassium hydroxide
The potassium hydroxide (KOH) test is a popular method used
to confirm onychomycosis, with accuracy dependent on proper
specimen collection, preparation, and examiner experience.
Specimen obtained from the nail bed and undersurface of the
nail plate is allowed to dissolve in 10% KOH solution after
placement on a glass slide for examination by light microscopy
(Figure 1).10 The test is inexpensive,15 and the results are
quickly available.1,10 The simplicity of the test enables it to be
performed in a clinician’s office.10 This test allows the examiner
to observe septate hyphae indicating that the patient is infected
with a dermatophyte, but no definitive information on the causa-
tive agent (i.e., genus and species) is obtained, and fungal via-
bility cannot be determined.10,15 As such, positive results from
KOH tests can be misleading if used as a test of cure.
Inconsistent sensitivity has also been associated with KOH
preparations.10,15 Sensitivity can be affected by a number of dif-
ferent factors, including the experience of the examining clini-
cian/technical staff, the preparation of the sample, and
differences in how samples are obtained. Moreover, a high rate
of false-negatives, ranging between 5% and 15%, has also
been reported because of low visibility and sparse distribution
of hyphae on the slide.5,16
Calcofluor white
Calcofluor white is a fluorescent agent that is mixed with KOH
in order to stain the chitin in the fungal cell wall, thus making
fungal elements more easily visible against the background of
host cellular material; however, as with KOH, the identity and
viability of the infecting microorganisms cannot be determined.17
Calcofluor white binds to the beta 1-3 and beta 1-4 polysaccha-
rides in cellulose and chitin and fluoresces when exposed to UV
radiation. Peak excitation and emission wavelengths for
calcofluor white solution are 365 and 435 nm, respectively
(Figure 1).10,18 The sensitivity issue associated with the
ª 2017 The International Society of Dermatology
Ghannoum et al.
(a)
Figure 1 Microscopic images of fungal
hyphae visualized using varying diagnostic
techniques. (a) Potassium hydroxide test
under a light microscope (9400
magnification). (b) Calcofluor white test
under a fluorescent microscope. (c)
Perdiodic acid–Schiff stain (9400 (c)
magnification). (d) Grocott’s methenamine
silver stain.10,14,41 Reprinted from Clinics in
Podiatric Medicine and Surgery, 2004;21
(4):565-578, Alberhasky RC, Laboratory
diagnosis of onychomycosis; and Clinics in
Dermatology, 2013;31:540-543, Gupta A
and Simpson FC, Diagnosing
onychomycosis, with permission from
Elsevier
traditional KOH test is overcome using calcofluor white if a
proper specimen is collected, although both techniques have
shown similar effectiveness.18 However, the significant barrier
to the regular use of this technique in office practice remains
the need for a fluorescence microscope.18
Fungal culture-based methods
Fungal culture has been considered to be the ‘gold standard’
technique in the diagnosis of onychomycosis.10,14 Clinical sam-
ples are plated onto a properly selected general media such as
potato dextrose agar with added antibiotics to inhibit overgrowth
by bacterial contaminants and a selective media such as Myco-
sel containing cycloheximide, which inhibits the growth of sapro-
phytic fungi.10 Using a curette to obtain adequate subungual
debris or multiple smaller fragments of nail plate and nail bed
has been suggested to improve the chances of obtaining a posi-
tive result. Distal nail clippings should not be submitted for fun-
gal culture because they often carry bacterial or saprophytic
mold contaminants that can easily overgrow dermatophytes on
culture media. Although the cultures are then incubated for at
least 4 weeks at 30 °C before being considered negative, the
culture could become positive for a dermatophyte within a
week. A significant advantage to using fungal culture is that it is
able to identify the causative agent,5 thus being more specific
than KOH testing.15
However, saprophytic mold or bacterial overgrowth may
occur, and there is a relatively long delay (e.g., 2–4 weeks)
associated with obtaining the results, which may lead to frustra-
tion for clinicians and their patients.10,15 The sensitivity of fungal
culture is also lower than a properly performed microscopy
test,10,19 and false-positive or false-negative results reduce the
reliability.
False negatives can occur for a number of reasons, including
insufficient nail material or improper sample collection too far
ª 2017 The International Society of Dermatology
Diagnosing onychomycosis by laboratory testing Review 3
(b)
(d)
removed from the site of infection.20 In order to reduce the like-
lihood of obtaining false-negative results, the specimen should
be collected as proximally to the infection as possible – an area
most likely to contain viable organisms and least likely to con-
tain contaminants.5 False-positive results may very rarely be
caused by contamination from organisms present as transient
flora or contamination of growth media.16
Histopathology
Histopathological techniques use direct microscopic examination
of tissue sections that have been stained with specific dyes to
visualize fungal growth patterns, which may suggest the pres-
ence of a dermatophyte.10,14 They cannot, however, determine
fungal viability.10
Periodic acid–Schiff stain
The periodic acid–Schiff (PAS) stain is a histopathology stain of
fungal polysaccharides performed on a nail plate biopsy speci-
men. It is highly sensitive and, when coupled with fungal cul-
ture, increases the overall sensitivity to 96%.1,15,21 The sample
is taken using a nail clipper which removes as proximally as
possible the full thickness nail plate and hyperkeratotic nail bed
if the latter is easily accessible. The sample is placed in forma-
lin and subsequently sectioned and stained with PAS stain,
which reacts with the aldehyde groups in the fungal cell walls to
produce a magenta-colored stain (Fig. 1).10 PAS staining
allows spores, hyphae, pseudohyphae, and yeast to be visual-
ized. In addition to high sensitivity, PAS staining results are
available quickly, usually within 24 to 48 hours.10 However, as
with KOH testing, the main disadvantages of PAS staining are
that the specific causative agent is unable to be confirmed and
that viable and nonviable organisms are indistinguishable.15
Furthermore, the cost associated with PAS is often higher than
other diagnostic techniques.22
International Journal of Dermatology 2017
4 Review Diagnosing onychomycosis by laboratory testing
Gomori’s methenamine silver
Gomori’s methenamine silver (GMS) staining is another com-
mon histopathology technique. The principles behind GMS
staining are similar to PAS staining, as the technique is also
based on the acid oxidation of fungal cell walls to aldehyde
10
groups. In GMS staining, chromic acid oxidizes polysaccha-
rides in fungal cell walls to aldehydes and reduces methena-
mine silver nitrate to metallic silver. The resultant stain shows
dark brown-colored cell walls on a pale green background
14
(Fig. 1). GMS staining is the most sensitive technique used to
identify dermatophytes and has been shown to be superior to
PAS staining.21 In one study, GMS stains detected an additional
23
five cases of onychomycosis from 51 PAS-negative stains.
Furthermore, GMS stains were shown to be qualitatively supe-
rior to PAS stains. However, both PAS and GMS staining tech-
niques expose laboratory staff to such chemicals as formalin
and phenol, which are known carcinogens and acute skin irri-
tants that should be handled in a chemical hood.
Molecular assays
Common targets and technologies
The ability of polymerase chain reaction (PCR) to amplify min-
ute amounts of target DNA from cells renders approaches
based on this technique particularly attractive for identification
of dermatophytes. There are many new molecular approaches,
including nested and semi-nested PCR (targeting specific genes
such as subtilisin, DNA topoisomerase II, and chitin synthase),
restriction fragment length polymorphism (RFLP) analysis, ran-
dom amplification of polymorphic DNA (RAPD), Southern blot
hybridization, electrophoretic mutation scanning, PCR-ELISA,
PCR-reverse line blot, sequence analyses (rDNA, rRNA
regions, and mnSOD), cDNA representational difference analy-
sis, oligonucleotide array, multiplex qPCR, and loop-mediated
isothermal amplification (LAMP). In addition, non-PCR
approaches or methods that combine PCR with other technolo-
gies, such as mass spectrometry, have been utilized. In this
article, we will discuss only those that have been used to detect
dermatophyte fungi directly from clinical samples.
PCR amplification of target genes
In one of the first studies to report the use of molecular tech-
niques to identify dermatophytes, Bock et al.24 employed a
PCR-based approach using a primer set targeting a fragment of
the gene coding for the fungal small ribosomal subunit 18S
rRNA. The DNA of seven dermatophytes (T. rubrum, T. menta-
grophytes, T. verrucosum, T. terrestre, M. canis, M. gypseum,
and E. floccosum) was amplifiable by these primers but not
DNA from 42 normal human skin samples. In addition, these
investigators collected 69 routine skin and nail specimens and
showed that PCR was more sensitive than culture in detecting
dermatophytes. Among 38 positive specimens, 35 were
detected by PCR, while only 28 were detected by culture
International Journal of Dermatology 2017
Ghannoum et al.
method, demonstrating the potential clinical relevance of a
PCR-based approach.
Baek et al.25 evaluated the sensitivity and specificity of cul-
ture-independent PCR amplification of fungal DNA in differenti-
ating between fungal species causing onychomycosis. These
investigators extracted DNA from both the nail plate and subun-
gual debris. Primers TR1 and TR2 amplified a 581-bp region of
the 18S rRNA for all fungi tested. Amplification of template DNA
using different primers generated specific fragments of the 18S
rRNA gene of fungi. Digestion of the amplicons with the restric-
tion endonuclease HaeIII resulted in a characteristic band pat-
tern and allowed differentiation of T. rubrum isolates. Combined
digestion with two or more restriction enzymes allowed differen-
tiation between dermatophytes, yeasts, and molds.
PCR-ELISA
A recent modification of the PCR approach is the combination
of PCR with enzyme-linked immunosorbent assay (PCR-
ELISA), which facilitates rapid identification of species-specific
DNA segments directly from clinical samples. Isolated genomic
DNA of nails is amplified with species-specific primers, with
PCR products subsequently detected using biotin-labeled
probes. Beifuss et al.26 recently reported using the PCR-ELISA
method for detection (within 24 hours) of the five common der-
matophytes, T. rubrum, T. interdigitale, T. violaceum, M. canis,
and E. floccosum, from clinical specimens. Genomic DNA was
isolated from skin and nail samples from patients with sus-
pected dermatophyte infections and was amplified with species-
specific digoxigenin-labeled primers targeting the topoisomerase
II gene. PCR-ELISA was performed using 204 microscopy-
positive samples in two university mycological laboratories in
Munich and Tubingen, and 316 consecutive specimens –
regardless of mycological findings – in a dermatological practice
laboratory. PCR-ELISA was confirmatory for one of the five der-
matophytes in 79.9% (163/204) of the clinical samples found to
be positive using microscopy.26 Cultures were positive for der-
matophytes in 59.8% of the same cases. A statistically signifi-
cant difference between these two methods for dermatophyte
detection was demonstrated.26 These findings suggest that
direct DNA isolation from clinical specimens coupled with PCR-
ELISA could provide a rapid, reproducible, and sensitive tool for
detection and discrimination of five major dermatophytes at
species level, independent of morphological and biochemical
characteristics.
Multiplex real-time PCR detection/identification
Arabatzis et al.27 used gene sequences to design a real-time
PCR assay for rapid detection and identification of dermato-
phytes in clinical specimens. Two assays based on amplification
of ribosomal ITS regions and employing probes specific to rele-
vant species and species-complexes were designed, optimized,
and clinically evaluated. The protocol was clinically evaluated
over 6 months by testing 92 skin, nail, and hair specimens from
ª 2017 The International Society of Dermatology
Ghannoum et al.
67 patients with suspected dermatophytosis. Real-time PCR
detected and correctly identified the causative agent in speci-
mens from which T. rubrum, T. interdigitale, M. audouinii, or
T. violaceum were cultured and also identified a dermatophyte
species in an additional seven specimens that were negative by
microscopy and culture.27 This highly sensitive assay also
proved to have high positive and negative predictive values
(95.7 and 100%, respectively), facilitating the accurate, rapid
diagnosis conducive to targeted rather than empirical therapy
for dermatophytosis.
PCR-reverse line blot
In a recent study, Bergmans et al.28 developed and successfully
used a PCR-reverse line blot (PCR-RLB) for rapid detection
and identification of nine dermatophyte species in nail, skin, and
hair samples. The developed method was based on ITS1
sequences using genus and species-specific probes in isolates
obtained from 819 clinical samples (596 nail, 203 skin, and 20
hair). In this method, membranes containing immobilized
oligonucleotide probes were exposed to denatured PCR prod-
ucts, allowed to hybridize for 30 minutes, then subjected to
stringency washes and detection using streptavidin-peroxidase
and chemiluminescence. The investigators reported a positive
PCR-RLB reaction in 93.6% of 172 culture-positive and micro-
28
scopy-positive samples.
MALDI-TOF
Matrix-assisted laser desorption ionization–time of flight
(MALDI-TOF) mass spectrometry (MS) is being adapted for use
in microbiology laboratories. The ability to analyze large biologi-
cal molecules by MS is made possible by the application of soft
ionization techniques that generate a spectrum of components.
In a MALDI-TOF analysis, a saturated solution of an organic
compound, or matrix, is added to a portion of a fungal colony,
and the mixture is then applied to a metal plate. Upon drying,
the crystallized mixture is subsequently irradiated using a laser
beam to force sublimation into a gas phase, followed by ioniza-
tion of the fungal sample.29
Ionized proteins within the sample are analyzed by a mass
spectrometer analyzer to produce a spectrum of mass-to-charge
32,37
Table 1 Diagnostic Tests for Onychomycosis.
Sensitivity Specificity
Test (%) (%)
Visuala 77 47
KOH 67–93 38–78
Culture 31–59 83–100
Periodic acid–Schiff 92 72
stain
PCR 95 100
a
Culture and periodic acid–Schiff staining.
ª 2017 The International Society of Dermatology
Diagnosing onychomycosis by laboratory testing Review 5
(m/z) ratios. These ratios measure how quickly charged ions
from the fungal sample move through the time of flight (TOF)
tube. Once spectra are generated, comparison of the m/z ratios
to a reference database leads to fungal identification. Protein
compositions differ between fungal species, which allows for
discrimination between closely related organisms.29
Several researchers have reported good correlation between
dermatophyte identification by conventional or gene sequencing
and MALDI-TOF analysis.30–32 However, all have had to supple-
ment current commercial databases with data generated from
their own reference dermatophyte strains. Alshawa et al.31
reported a correlation rate of 91.9% of dermatophyte strains
using conventional methods, while Jensen et al.33 matched
99.3% of identifications derived by gene sequencing. Clearly,
MALDI-TOF will prove an invaluable methodology for rapid
identification of dermatophyte isolates once commercial data-
bases become more robust. However, another current limitation
of this method is the necessity for isolating pure dermatophyte
colonies from clinical samples before MALDI-TOF assay can be
attempted. Further research to enable detection of dermato-
phytes directly from clinical samples is required.
Mixed testing results
The frequency of inconsistent results (i.e., opposing results from
different diagnostic tests) remains a challenge in onychomyco-
sis diagnosis (Table 1). A study conducted by Clayton et al.
showed 11% of 2113 toenails were KOH positive but fungal cul-
ture negative.19 A negative result obtained from fungal culture
can be misleading, as it has been reported that almost half of
all onychomycosis samples fail to yield a positive culture.5 Prob-
lems with obtaining a suitable sample contribute to the high
false-negative rate for fungal culture, with results as high as
60%.34 Despite this, it is widely regarded that onychomycosis is
confirmed most accurately by positive results from fungal cul-
ture.5 In addition, false-positive results from KOH tests do not
ascertain that the observed fungi are viable and may accurately
reflect the state of the infection. Examples of differences
between KOH and culture results have been observed in many
clinical trials of new antifungals being developed to treat
Confirm Viability of Identification of Cost of Time for
Fungi Organism Testing result
No No $
No No $ 5–30 min
Yes Yes $$ Up to 4 weeks
No Yes $$ 24–48 h
No Yes $ Within 24 h
International Journal of Dermatology 2017
6 Review Diagnosing onychomycosis by laboratory testing
onychomycosis. For example, in a clinical trial evaluating tava-
borole, 5% topical solution, it was observed that the positive
KOH test results were notably higher than positive culture
results (68.8–62.9% vs. 14.6–13.0%, respectively), presumably
because dead fungal elements were detected from the residual
nail specimen.35 This is just one example of the importance of
properly obtaining a suitable nail specimen and its preparation
to permit growth in a culture medium15 and the importance of
using a sensitive technique (e.g., calcofluor white staining) and
having an experienced person to read the direct examination.
The importance of confirmatory testing
Prior to prescribing an antifungal treatment for a suspected fun-
gal infection, the American Academy of Dermatology recom-
mends the positive identification of a fungal infection (i.e.,
positive mycology).36 This confirmed diagnosis of onychomyco-
sis and the correct identification of the causative agent are
imperative, as treatment can be lengthy and costly. Further-
more, a misdiagnosis may expose patients to the wrong therapy
or impact the patient’s perception of therapeutic effective-
ness.4,36 Performing confirmatory testing before the initiation of
treatment is associated with substantial savings as compared to
costs involved when treating empirically.37
The treatment approach for onychomycosis is dependent on
the severity of the infection, degree of involvement of the nail
plate, and medical history of the patient. Oral medications are
administered over a relatively short treatment period (e.g.,
12 weeks) and have high mycological and clinical cure rates
(i.e., clinically normal nail with negative mycology), but they are
associated with systemic adverse effects and possible drug
38
interactions. Topical medications are recommended for use in
mild-to-moderate infection and are associated with fewer
adverse events but have a longer treatment period.
Obtaining a correct diagnosis and mycological identification
of the causative agent will inform the course of action for treat-
ment. For example, it has been reported that nondermatophyte
molds are resistant to the usual antifungal treatments pre-
scribed for onychomycosis, and thus a confirmatory diagnosis
of a nondermatophyte pathogen such as Fusarium or Candida
albicans would be crucial to prescribing alternative treat-
ment.29,39 Therefore, confirmatory testing would dictate the use
of a drug that is effective against both dermatophytes and non-
dermatophyte molds. For example, a recent case report involv-
ing Paecilomyces lilacinus-induced onychomycosis – which is
extremely difficult to treat – was identified through proper labo-
ratory technique and treated with the topical combination of
40
tavaborole and efinaconazole.
Patients who have undergone a treatment plan directed at
addressing the specific causative pathogen are more likely to
have a better outcome, including a better quality of life because
of resolution of the infection, than patients who received untar-
geted treatment.
International Journal of Dermatology 2017
Ghannoum et al.
Conclusions
Distinguishing onychomycosis from other disorders such as
psoriasis can sometimes be challenging for clinicians based on
physical examination alone. Despite the availability of a variety
of laboratory and diagnostic techniques for onychomycosis, no
single routinely available technique is both highly specific and
sensitive, able to identify the causative pathogen, and able to
provide rapid results at low cost. The diagnosis of onychomy-
cosis could therefore benefit from improvements to existing
techniques in order to ensure improvement in the quality of
results. Alternatively, new techniques could be developed with
the aim of combining the advantages of several individual
techniques. At present, the fluorescent calcofluor stain or nail
plate biopsy with PAS testing combined with fungal culture are
most commonly utilized as the most acceptable and practical
means of confirming the diagnosis of onychomycosis in clinical
laboratories, although MALDI-TOF mass spectrometry is
becoming more widely used. Currently, the cost of PCR is the
limiting factor in its routine use for onychomycosis diagnosis
within general practice. Although direct examination by 10%
KOH is often performed as a preliminary test at the physi-
cian’s office, it is recommended that the entire sample be sent
to the referring laboratory in order to ensure the best chance
of obtaining both positive microscopy and positive fungal cul-
ture. Optimal management of onychomycosis can be costly,
requires highly skilled personnel, and requires management
over a lengthy period.
Questions (answers provided after
references)
1 Why do some people develop onychomycosis while others do
not when exposed to the same environmental factors?
a. Differences in personal hygiene
b. Genetic predisposition
c. Wearing sandals or open-toed footwear
d. All of the above
2 What inflammatory disorders can mimic onychomycosis?
a. Rheumatoid arthritis
b. Psoriasis
c. Lichen planus
d. Crohn’s disease
3 Why is it important to clean the nail before collecting a
sample?
a. It softens the nail for easier cutting.
b. It protects the hands of clinical staff from contamination.
c. It removes contamination bacteria and saprophytic molds.
d. It protects the patient from possible infection during collec-
tion procedures.
4 Microscopy is a more specific test for dermatophytes than
culture. True or False?
ª 2017 The International Society of Dermatology
Ghannoum et al.
5 Why is it important not to use microscopy alone for a test of
cure?
a. The hyphae may be nonviable, leading to a false-positive
diagnosis.
b. Fungal species cannot be identified by direct microscopy
of subungual debris.
c. It is too expensive and time consuming.
d. The procedure needs to be done by experienced person-
nel.
6 What is the advantage of identifying the infecting organism in
a case of onychomycosis?
a. It aids in the diagnosis of superficial white onychomycosis.
b. It will identify the environmental source of the infection.
c. It is often required for insurance coverage.
d. It will help in determining proper antifungal treatment.
7 What are the two disadvantages of PAS and GMS staining
techniques?
a. Cost of the procedure
b. Length of time required to obtain results
c. Exposure of technicians to dangerous chemicals
d. Necessity for performance of biopsy.
8 What is the rationale behind developing molecular assays for
identifying dermatophytes in onychomycosis cases?
a. Greater sensitivity
b. Greater specificity
c. Lower cost
d. All of the above.
9 What is the main pitfall in using PCR to identify dermato-
phytes directly from nail specimens?
a. DNA from contaminating fungi is also amplified.
b. Testing must be done on biopsied samples.
c. PCR cannot identify fungi to genus/species.
d. Fungi that cause nail infections are not included in com-
mercial databases.
10 What is the current limitation of the MALDI-TOF assay for
identifying dermatophyte strains?
a. It cannot be performed on clinical samples.
b. Dermatophyte Identifications do not correlate well with
PCR results.
c. Commercial databases are inadequate.
d. Cost per test is extravagant.
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