http://informahealthcare.

com/hth
ISSN: 0265-6736 (print), 1464-5157 (electronic)

Int J Hyperthermia, 2013; 29(6): 544–550

! 2013 Informa UK Ltd. DOI: 10.3109/02656736.2013.823672

RESEARCH ARTICLE

Antifungal efficacy of lasers against dermatophytes and yeasts in vitro

Uwe Paasch1, Andrea Mock1, Sonja Grunewald1, Marc O. Bodendorf2, Michael Kendler1, Anna-Theresa Seitz1,
Jan C. Simon1, & Pietro Nenoff3
1
Klinik und Poliklinik für Dermatologie, Venerologie und Allergologie, Abteilung Laser und Kosmetik, Universitätsklinikum Leipzig Anstalt
öffentlichen Rechts, 2Practice for Dermatology and Pathology Doctors Reusch und Mielke, Hamburg, and 3Labor für medizinische Mikrobiologie,
Partnerschaft Professor Pietro Nenoff und Dr Constanze Krüger, Mölbis, Germany

Abstract Keywords
Purpose: Approximately 2–13% of the world population suffers from onychomycosis. Recently, Dermatology, dermatophytes, fungi, laser,
lasers have been introduced for treatment. However, no effect was found with in vitro laser nails, skin, yeasts
irradiation of pathogens on agar plates. This study aimed to investigate the efficacy of laser
irradiation against fungi using an alternative in vitro approach. History
Materials and methods: Lasers of 808, 980 and 1064 nm were used to heat cell culture media
and a nail clipping. Trichophyton rubrum, T. interdigitale, Microsporum gypseum, Candida Received 8 March 2013
albicans, C. parapsilosis, and C. guilliermondii species were subcultured and subjected to laser Revised 6 June 2013
treatments (808/980 nm: 9–27 J/cm2, 6 ms, 12  12 or 12  50 mm and 1064 nm: 50–240 J/cm2, Accepted 5 July 2013
90 ms, 5–10 mm). After irradiation, the fungal elements were transferred onto agar plates using Published online 19 August 2013
conventional and Drigalski spatulas and were incubated for 6 days.
Results: The highest increase in temperature was found using a 980-nm laser with a pulse
duration of 6 ms and a fluence of 27 J/cm2. The histology work-up revealed a dissection of the
nail plate from the nail bed tissue after laser irradiation. Growth inhibition was only found for
C. guilliermondii and T. interdigitale. All other pathogens presented only reduced growth, and
C. albicans growth was unaffected.
Conclusions: This study demonstrates a clear thermal effect for linear scanning 980-nm and
long-pulsed 1064-nm laser systems on either nail clippings or cell culture media. Complete
pathogen growth impairment was achieved if temperatures were measured above 50  C. The
results for the 1064-nm system were almost comparable to 980 nm results.

Introduction contact with hyphal fragments and arthroconidia. Household

dust enables the preservation of spores for years [2].
Dermatophytes are a group of filamentous pathological fungi
The typical clinical diagnosis is confirmed by mycological
that can inhabit human skin, hair and nail plates. They are
examination using direct microscopy with a potassium
ubiquitous in distribution and produce superficial infections
hydroxide preparation, cultivation on Sabouraud’s dextrose
called dermatophytosis [1]. Dermatophytosis is found in
agar, and molecular methods such as PCR (polymerase chain
approximately 20–25% of the world population [2]. An
reaction) [6,11]. However, because of the high frequency of
estimated 2–13% of the population suffers from onychomy-
cosis (OM) [3–5], which is the most common disease of nails
potential false negative laboratory results accompanying 13
20
typical clinical symptoms, antifungal treatment is advisable
worldwide and is responsible for approximately half of all nail
[4]. Topical antifungal formulations are available but are only
abnormalities [6]. The condition produces a huge impact on
helpful in distal subungual onychomycosis cases affecting up
quality of life [7,8]. The primary age group affected consists
to one third of the nail plate. Systemic treatment in
of elderly patients.
combination with topical therapeutics is advisable if larger
Frequently, dermatophytes are causative agents for OM in
areas of the nails or more than 3 out of 10 nails are affected
temperate western countries [2,9]. These fungi include the
[12]. Despite a variety of available compounds (e.g. terbina-
genera Epidermophyton, Microsporum and Trichophyton [1].
fine, fluconazole, and itraconazole), standard systemic ter-
Among these fungi, T. rubrum and T. interdigitale (formerly
binafine administration achieves a disease-free nail in only
T. mentagrophytes) are the most often (90–93%) detected in
approximately 35–76% of cases [13–15]. In addition, up to
OM [2,6,10]. The transmission of these fungi occurs by
22.3% relapse rates are found within 3 years after
completed treatment [16]. Furthermore, in geriatric patient
Correspondence: Prof Dr med. habil. Uwe Paasch, MD, Klinik und cohorts, the regularity and efficacy of topical application is
Poliklinik für Dermatologie, Venerologie und Allergologie, Abteilung often reduced, and systemic medications may affect the liver
Laser und Kosmetik, Universitätsklinikum Leipzig, Philipp Rosenthal
Strabe 23, 04103 Leipzig, Germany. Tel: +49-341-9718611. Fax: +49- or interfere with other drugs [17]. Therefore, the long-term
341-9718619. E-mail: uwe.paasch@medizin.uni-leipzig.de outcome of systemic treatments is unsatisfactory because of
DOI: 10.3109/02656736.2013.823672 Lasers to treat fungal nail infections 545

97% of treated patients with improvements accord-

mycosis using 870- and 930-nm light exposure:

therapeutic failure, relapse, reinfection, and the resulting

85% ‘clear nail linear extent.’ Disposable auto-

Treatment of mild, moderate, and severe onycho-
adverse events [18].
Studies of alternative treatment options such as surgery

FDA USA 510 k:K103626 Thermal sensor

79.6% positive response after 2 treatments

Table I. Common laser systems available for treatment of onychomycosis. Applied laser parameters for the treatment of onychomycosis using a Nd:YAG laser and the laser parameter range used.
[19], high frequency waves [20], photodynamic therapy

FDA USA 510 K: K093547 K093545

[21,22], and CO2 laser-based therapy [23] yielded discoura-

Reference/comments
ging results. The huge amount of heat applied with early CO2
lasers was helpful in managing hyperkeratotic conditions such

FDA USA 510 K: K0110370

FDA USA 510 K: K111483

FDA USA 510 K: K103338

FDA USA 510 K: K113810

ing to the manufacturer
as onychogryphosis [24] and in performing matricectomies

mated toe pod system.

FDA USA 510 k:071815
[23,25]. In contrast, the rather cold ablation effect of 2940-nm
Er:YAG lasers has been shown to be unsuitable for the
inhibition of fungal growth [26]. In addition, the use of

Thermal sensor

Clear Choice
intense pulsed light (695–1000 nm), flashlamp pulsed dye

White paper
Clear Sense

Clear Steps
laser (585 nm) and the KTP laser (532 nm) have also been

[33, 36]

20 Hz
shown to be ineffective [26]. As CO2 lasers were found to be

[32]
effective but unpredictable, q-switched [26] and longer pulsed
near-infrared lasers [27,28] with no ablation should have a
much better side-effect profile while maintaining their

CoolBreeze/CoolTouch Varia, CoolTouch

efficacy. It has been postulated that multiplexed wavelengths

Noveon/Nomir Technologies Naillaser,

of 870 and 930 nm produce their lethal effects against fungi

System name Manufacturer

via a decrease in mitochondrial membrane potential together
with a simultaneous increase of ROS (reactive oxygen

Joule, Clear Sense Sciton

species) [29]. Operating at physiological temperatures, a

Medical Technologies

Aerolase, Light pod Neo

clinical improvement was observed in OM after four treat-

Genesis Plus, Cutera

AccuSculpt Lutronic
Pin Pointe Cynosure
Primogent Solutions

Q-Clear, Light Age

ments [30]. To destroy fungi in a highly selective way, an 800-

SP Dualis, Fotona

Alma Harmony
nm femtosecond Ti:sapphire laser has been used to apply
200  1015 s pulses at 76 MHz pulse frequencies. In add-
ition, 7  1031 photons m2 s1 intensities or above have Lunula
successfully eliminated fungi in nail clippings after a single
treatment, and this has been managed without any destruction
of the nail plate at up to intensities of 1.7  1032 photons
m2 s1 [31]. Because femtosecond lasers are not easily

1–1.5 defocused up to 6 mm
available, and lasers operating at a 1064-nm wavelength are
Spot size (mm)

widely used for other indications, the latter type of laser has
also been investigated with respect to its suitability for the
(2862)

1–1.5

2.5–6
3–10
n.a.

treatment of nail fungi. Nd:YAG systems at this wavelength

3
1
4
have been shown to be effective if operated in a q-switched
manner [26]. Using a 0.65-ms pulse duration while applying a
2-mm spot with the energy fluence set at 223 J/cm2 in the
absence of any cooling sprays, gels or topical anesthetics
produces an 87.5% clearing rate after three treatments every
3 weeks [32]. In clinical studies it has been shown that a rise
Pulse duration (ms)

of temperature within the nail plate over a given time might

12 min per foot

0.003–0.01
0.3–200

be crucial for the clearing of onychomycosis. Using a 1064- 0.1–0.3

0.1–1

0.65
0.45
0.45

0.02
n.a.

n.a.

0.1
25

nm Nd:YAG laser set to a pulse duration of 35 ms to deliver

35–40 J/cm2, 45  5  C was achieved at the nail plate after
being irradiated three times. The procedure was repeated four
times weekly, resulting in a 95.8% fungal clearing rate [33].
However, to date, many systems are available that operate at
500–1200 mJ/pulse
Fluence (J/cm2)

rather diverse parameter settings, although most often, the

0.95 per diode

20–450 J/cm2
250 mJ/Puls
212/414

1064-nm wavelength is chosen (Table I). Recently, the

35–40
5200

5–40

4–12
25.5
223

efficacy of a 1064/532-nm Nd:YAG-system was tested

in vitro using q-switched pulses as well as long pulses
(1064 nm only) on T. rubrum. Unfortunately, no effect was
observed from the irradiation of pathogens on agar plates
after incubation at 30  C for 3–6 days [34]. However, the
n.a., not applicable.
Wavelength (nm)

experimental design might not have been ideal, and contrast-

532 qs þ 1064
long pulsed

ing findings might have been found.

As fungal infection of the human skin and its appendages
405/635

870/930

1064 qs

are so common and difficult to treat on both sides, and a

1064

1064

1064

1064
1064
1320

1440

variety of laser systems using different wavelengths and

546 U. Paasch et al.
settings that have been shown to be effective against causative
fungi, this study aimed to investigate the efficacy of laser
irradiation in vitro against common pathogens using an
alternative approach of pathogen culture post-irradiation.
Materials and methods
The objective of this study was to define the impact of 808-,
980-, and 1064-nm lasers using various parameter settings to
deliver heat to cell culture media and to destroy common
species and different strains of fungi in in vitro culture
conditions.
Temperature measurements
Six-well ELISA reader plates (83.1839.300, Sarstedt,
Newton, NC, USA) were used for irradiation of pure cell
culture media to measure the heating effect of the tested laser
systems. The temperature was evaluated by replicate meas-
urement using a contact-free laser thermometer (MiniTemp4,
Raytek, Berlin, Germany).
Pathogens
T. rubrum, T. interdigitale, M. gypseum, C. albicans,
C. parapsilosis, and C. guilliermondii species serving as
reference pathogens at the laboratory for medical microbiol-
ogy were taken and subcultured into a Sabouraud’s broth
before being further subjected to laser treatment. Although
T. rubrum and T. interdigitale are the most common
pathogens, the intentions to integrate the other pathogens
were to see if there are possible differences in heat
susceptibility. Liquid cultures of those pathogens were
incubated at 37  C and 27  C, respectively, for 3 days to a
maximum of 12 days until the Sabouraud’s broth became
murky and clouded as an optical indicator of fungal growth.
In an initial pathogen eradication experiment, 2 mL of the
fungal broth was transferred into glass Petri dishes or each
well of the 6-well plastic ELISA plates for further laser
treatment. To achieve a clinically relevant temperature
scenario the samples were kept at room temperature before
processing. After laser irradiation, the fungal broth samples
were transferred in three sections onto agar plates (Figure 1).
For the third experiment, 10 mL and 50 mL of Sabouraud’s
broth were transferred after laser treatment onto agar plates
using Drigalski spatulas. Next, 25 mL of the same Sabouraud’s
broth served as a control. The colonies were counted on day 6
post treatment.
Laser treatment
Laser treatment was performed using three different systems:
an 808-nm linear scanning diode laser, a 980-nm linear
scanning diode laser, and a long pulsed 1064-nm Nd:YAG-
laser (all Alma Lasers, formerly Quantel-Derma and
Wavelight, Erlangen, Germany). The latter is mainly
indicated for hair removal, vascular lesion treatments and
achieving non-ablative skin rejuvenation, and the first two
systems are routinely used for hair removal.
Laser treatment was performed in three experiments: (1)
the evaluation of the ability of a 980-nm laser to heat water
together with the evaluation of 808 and 980 nm linear
Int J Hyperthermia, 2013; 29(6): 544–550
scanning diode laser to heat fungi in media (Sabouraud’s
broth, Figure 2), (2) the testing of the ability of the 980-nm
linear scanning diode laser as well as a long pulsed 1064-nm
Nd:YAG-laser to efficiently destroy the entire range of
pathogens of interest at the laser parameters with an improved
read out, and (3) to evaluate the 1064 nm effect on nail
clippings in vitro.
The parameter settings used in the latter two experiments
were selected according to the results of the first experiment:
808 nm, a fluence of 9–27 J/cm2 and a pulse duration of 6 ms
at a spot size of 12  12 (6-well plates) or 12  50 mm (glass
Petri dishes); 980 nm, a fluence of 9–27 J/cm2 and a pulse
duration of 6 ms at a spot size of 12  12 (6-well plates) or
12  50 mm (glass Petri dishes); and 1064 nm, a fluence of
50–240 J/cm2, a pulse duration of 90 ms and a varying spot
size of 5 to 10 mm. All experiments were conducted using
glass Petri dishes or plastic 6-well ELISA plates.
Histology
A routine histology work-up was performed on the nail
clippings to evaluate the effect of the 1064-nm Nd:YAG laser.
After removing the nail parts, because of other indications
and ensuring informed consent from the patient, the nail parts
were subjected to laser irradiation using a fluence of 70 J/cm2
and a pulse duration of 40 ms (three passes). The tissue block
was embedded in paraffin, cut into 5–8-mm slices and stained
with haematoxylin and eosin (H&E) and periodic acid Schiff
(PAS) according to laboratory standard procedures. The
biopsies were evaluated under a calibrated microscope
(BX41, Olympus Germany, Hamburg) equipped with a digital
camera (DP70, Olympus Germany). The specimen was
measured using calibrated CellF software (Olympus
Germany).
Statistics
Statistical analysis was performed using Statistica 7.0 soft-
ware for Windows (StatSoft, Tulsa, OK, USA). A Mann-
Whitney U test was performed to investigate the differences
between the groups. P values 50.05 were considered statis-
tically significant.
Figure 1. Illustration of how the fungi were transferred to culture plates
after incubation.
DOI: 10.3109/02656736.2013.823672
Figure 2. Temperatures within water and cell culture media containing pathogens in response to repetitive laser treatments (stacks) at various
parameter settings.
Results
Heating of cell culture media
When using the 808-nm system at a pulse duration of 6 ms and
a fluence of 9 J/cm2, the temperature curve did not reach 22  C,
even after a long series of stacks were applied (Figure 1). A
temperature of 26  C could only be achieved if the fluence was
raised to 27 J/cm2 with 30 stacks. When the 980-nm system was
used, there was a slow increase in the temperature of cell
culture medium without pathogens when using a pulse duration
of 6 ms and a fluence of 9 J/cm2. The increase in temperature
was higher if pathogens were added to the cell culture medium.
The most pronounced increase in temperature was found if a
980-nm system was applied using a 6-ms pulse duration to
apply a fluence of 27 J/cm2. Overall, the time needed to heat
the medium with and without pathogens was clearly dependent
on the amount of stacks, and therefore exposure time at a given
wavelength, fluence and pulse duration. While the 808-nm
system did not produce sufficient heating at low fluences and
delayed at higher fluences, the 980-nm system was proven to
be efficient in a short time period. Higher fluences resulted in a
more rapid heating.
Efficacy of laser treatment in eradication of common
and rare medically important fungi
Efficacy of laser irradiation on fungal pathogens cultured in
liquid media
The 980-nm system was used to heat the fungi within the
culture media over time. During the application of a fluence
of 27 J/cm2 at a 6-ms pulse duration (318 pulses) during a
9-min time period, a temperature of approximately 45  C was
Lasers to treat fungal nail infections 547
reached at steady state using the 12  12 mm spot. Using the
12  50 mm spot resulted in a longer time period (21 min)
until a temperature steady state was achieved with 318 pulses.
After transferring the pathogens onto agar plates, at day 6
after treatment, the colonies were counted and growth
inhibition was found for only one run of C. guilliermondii
and T. interdigitale. All other runs demonstrated reduced
growth compared with untreated controls (C. guilliermondii,
C. parapsilosis, T. interdigitale, and M. gypseum). The growth
of the yeast C. albicans was never disturbed (Table II). The
results indicated that complete clearance was achieved only if
the measured temperatures exceeded 50  C. This is in line
with information that spores will survive temperatures up to
60  C [35].
The in vitro effect of the 1064-nm system was even worse.
Pathogen growth was inhibited only temporarily using high
fluences (Table III).
Histology
The routine histology work-up performed on nail clippings to
evaluate the effect of the 1064-nm long pulsed laser showed a
clear dissection of the nail plate from the nail bed, indicating
a heat action at the target tissues as known from other lasers
(Figure 3).
Discussion
Onychomycosis is a worldwide emerging problem with no
tendency for self-healing and the potential to affect surround-
ing tissues, as well as the possibility of predisposing the
patient to secondary bacterial infections. In conjunction with a
wide variety of topical but less effective treatment options,
548 U. Paasch et al. Int J Hyperthermia, 2013; 29(6): 544–550

Table II. Efficacy of 980-nm irradiation of pathogens in liquid culture media as measured 6 days post-
treatment on culture plates.

Pathogen LEDA Temperature Results 6 days post-treatment

2 
C. guilliermondii 27 J/cm , 6 ms 58 C Complete clearance
N¼5 12  12 mm
318 pulses, 9 min
27 J/cm2, 6 ms 37–45  C Reduced growth
12  12 mm
318 pulses, 9 min
C. albicans 27 J/cm2, 6 ms 48  C No inhibition
N¼5 12  50 mm
318 pulses, 21 min
27 J/cm2, 6 ms 45–47  C No inhibition
12  12 mm
318 pulses, 9 min
C. parapsilosis 27 J/cm2, 6 ms 58  C Reduced growth
N¼5 12  50 mm
318 pulses, 21 min
27 J/cm2, 6 ms 37–45.5  C Reduced growth
12  12 mm
318 pulses, 9 min
T. interdigitale 27 J/cm2, 6 ms 45.5  C Reduced growth
N¼7 12  12 mm
318 pulses, 9 min
27 J/cm2, 6 ms 55  C Complete clearance
12  50 mm
318 pulses, 21 min
27 J/cm2, 6 ms 41.5–46  C Reduced growth
12  12 mm
318 pulses, 9 min
M. gypseum 27 J/cm2, 6 ms 45.5–51  C Reduced growth
N¼5 12  12 mm
318 pulses, 9 min
T. rubrum 30–51 J/cm2 n.m. Reduced growth n ¼ 5
N¼5 12–18 ms Increased growth
12  12 mm n ¼ 1 (33 J/cm2, 12 ms)
40–100 pulses

N, the number of repetitions per strain.

Table III. Efficacy of 1064-nm irradiation of pathogens in liquid culture media as measured 6 days post-treatment on culture plates.

Pathogen Pulses 50 J/cm2 40 ms 100 J/cm2 40 ms 240 J/cm2 40 ms

C. guilliermondii
N¼5 2 No inhibition
C. albicans
N¼5 2 No inhibition
C. parapsilosis
N ¼ 5 240 J/cm2 2 Growth inhibition
N ¼ 5 100 J/cm2
N ¼ 5 50 J/cm2 15 Growth inhibition Growth inhibition
T. interdigitale
N ¼ 5 240 J/cm2 15 Temporary growth
inhibition day 1–2
N ¼ 5 100 J/cm2 12 Temporary growth
inhibition day 1–2
N ¼ 5 50 J/cm2 12 No inhibition
M. gypseum
N ¼ 5 240 J/cm2 15 Increased growth Increased growth Increased growth
N ¼ 5 100 J/cm2 12 Increased growth Increased growth Increased growth
N ¼ 5 50 J/cm2 12 Increased growth Increased growth Increased growth
T. rubrum
N ¼ 1 40 J/cm2, 15 ms 40 J/cm2 40 J/cm2 70 J/cm2 100 J/cm2 100 J/cm2
N ¼ 2 40 J/cm2, 30 ms 15 ms 30 ms 40 ms 65 ms 90 ms
N ¼ 1 70 J/cm2, 40 ms 100 Increased growth No reduced growth –
N ¼ 1 100 J/cm2, 65 ms 31 – Reduced growth – –
N ¼ 1 100 J/cm2, 90 ms 100 pulses, 3 Hz – No reduction or – –
increase in growth

N, the number of repetitions per strain.

DOI: 10.3109/02656736.2013.823672
Figure 3. Histology of a nail clipping after applying a 1064-nm long
pulsed Nd:YAG laser at a fluence of 70 J/cm2 and a pulse duration of
40 ms (three passes), PAS (Periodic acid Schiff).
standard systemic terbinafine administration achieves a
disease-free nail in only approximately 35–76% of cases
[13–15]. In addition, up to 22.3% relapse rates are found
within 3 years after the completion of treatment [16].
Recently, the option of laser nail fungus treatment has
appeared. This therapy modality seems to be especially
appropriate for the elderly patient. Typically, a decrease in
flexibility and an increase in co-medication are observed in
this cohort of patients. Thus, using a local therapy with no
systemic effects from drug interference or metabolic effects
would be beneficial for the elderly. To date, a variety of laser
systems have been promoted to treat onychomycosis, and
some of them already carry FDA approval for nail clearance.
Most often, 1064-nm systems are used, and superior clearing
rates from 87.5% [32] up to 95.8% [33,36] have been
published. However, the latter cure rates were published
without any peer review. In contrast, no effect at all has been
found by in vitro irradiation of pathogens on agar plates [34].
As fungal infection of the human skin and its appendages is so
common and a variety of laser systems using different
wavelengths and settings are available, this study aimed to
investigate laser irradiation efficacy against common and rare
pathogens in vitro by performing three in vitro experiments.
For the long pulsed near-infrared lasers, heat is the most
likely mechanism of action. Here, we clearly demonstrate that
lasers emitting at 808 nm and 980 nm are effective in heating
cell culture media with and without (neat cell culture
medium) pathogens. The use of higher wavelengths and
higher fluences are advantageous. However, with both there is
an increase in pain if applied clinically. Basically, these
results might direct us to use linear scanning hair removal
lasers for experimental treatment of fungal pathogens,
although clinically effective, safe and painless treatment
protocols still need to be established. In addition, this study
revealed that rather long stacking at a higher fluence is
required to heat up cell culture media even in small volumes.
Unfortunately, this might not really reflect the situation
in vivo where the pathogens are not kept in a liquid
environment. In contrast, a clinical treatment over a longer
time using more stacks might be beneficial for homogeneous
nail plate heating and to ensure definitive pathogen
Lasers to treat fungal nail infections 549
eradication since there is no specific absorber carried by the
fungi to be destroyed. In comparison, the application of a
1064-nm long pulsed Nd:YAG using a fluence of 70 J/cm2
and a pulse duration of 40 ms (three passes) resulted in a clear
dissection (clefting) of the nail plate from the nail bed,
indicating a heat action at the target area or tissues. Clefting is
often found at the junction between epidermis and dermis in
response to other laser interventions, indicating a cumulative
temperature effect. To what extent this effect is purely
coincidental or a necessity for effective treatment needs to be
further elaborated.
During the latter two experiments we tested the efficacy of
the lasers in destroying pathogens in vitro. Other researchers
have been unable to demonstrate the eradication of pathogens
by direct irradiation of agar plates [34]. Using our alternative
approach of transferring irradiated pathogens to agar plates it
was also demonstrated that lasers do not always have a
harmful effect on fungi. This was especially the case in the
two most common species T. rubrum and T. interdigitale.
Indeed, in some cases, growth appears to have been induced.
Most likely, the temperatures achieved were not high enough
or were unevenly distributed, which might result in an
occasional stimulation sometimes described as ‘low-level
laser stimulation’. With this taken into account, an on-time
temperature measurement might be helpful in evaluating the
effects of the lasers and temperature. Otherwise, it remains to
be proven if effective temperatures above 50  C are really
necessary, and if so, how they can be applied in vivo with no
or low pain.
Conclusion
Dermatophytosis is found in approximately 20–25% of the
worldwide population [2]. Out of the worldwide population, an
estimated 2–13% of the population suffers from onychomy-
cosis [3–5], which is the most common disease of nails
worldwide and constitutes approximately half of all nail
abnormalities [6]. The disease has a huge impact on quality of
life [7,8]. Recently, the option of laser nail fungus treatment
has appeared. The reported superior treatment effects of 87.5%
[32] up to 95.8% [33,36] clearance are in contrast to in vitro
studies where no effect has been found following the irradi-
ation of pathogens [34]. This study demonstrates a clear
thermal effect of linear scanning 808-, 980 - and long pulsed
1064-nm laser systems on cell culture media or nail clippings.
With shorter wavelengths, higher fluences and more stacks
were needed. However, the post-irradiation growth of fungi
was only occasionally inhibited. Finally, growth inhibition
appears to be strongly dependent on the temperature reached.
Most likely, homogeneous heat distribution is crucial, although
nothing is currently known about the minimal time needed for a
given plateau temperature. Further studies with on-time in vivo
temperature evaluation, e.g. during the treatment, and clinical
long-term follow-ups are needed to evaluate the in vivo laser
and thermal effect for each specific system.
Acknowledgements
The authors wish to thank Cornelia Schneider for her expert
technical assistance.
550 U. Paasch et al.
Declaration of interest
The 808-nm and 980-nm laser systems were loaned from the
manufacturer to run a separate hair removal study. Uwe
Paasch and Marc Bodendorf are working as an honorarium
speaker for Alma Lasers. Uwe Paasch and Sonja Grunewald
served as consultants for Quantel-Derma, now Alma Lasers.
Uwe Paasch and Jan C. Simon received unrestricted research
grants from Quantel-Derma, now Alma Lasers. The authors
alone are responsible for the content and writing of the paper.
References
1. Ameen M. Epidemiology of superficial fungal infections. Clin
Dermatol 2010;28:197–201.
2. Havlickova B, Czaika VA, Friedrich M. Epidemiological trends in
skin mycoses worldwide. Mycoses 2008;51:S2–15.
3. Hamnerius N, Berglund J, Faergemann J. Pedal dermatophyte
infection in psoriasis. Br J Dermatol 2004;150:1125–1128.
4. Amichai B, Davidovici B, Trau H, Lyakhovitsky A, Grunwald MH,
Shemer A. A rationale for systemic treatment in onychomycosis
with negative results on fungal examination. Clin Exp Dermatol
2011;36:724–727.
5. Scher RK, Tavakkol A, Sigurgeirsson B, Hay RJ, Joseph WS, Tosti
A, et al. Onychomycosis: Diagnosis and definition of cure. J Am
Acad Dermatol 2007;56:939–944.
6. Nenoff P, Ginter-Hanselmayer G, Tietz HJ. Onychomykose – ein
Update: Teil 1 – Prävalenz, Epidemiologie, disponierende Faktoren
und Differenzialdiagnose. [Fungal nail infections – an update: Part
1 – Prevalence, epidemiology, predisposing conditions, and differ-
ential diagnosis]. Hautarzt 2012;63:30–38.
7. Gupta AK, Jain HC, Lynde CW, Macdonald P, Cooper EA,
Summerbell RC. Prevalence and epidemiology of onychomycosis
in patients visiting physicians’ offices: A multicenter Canadian
survey of 15 000 patients. J Am Acad Dermatol 2000;43:244–248.
8. Elewski BE. Onychomycosis. Treatment, quality of life, and
economic issues. Am J Clin Dermatol 2000;1:19–26.
9. Elewski BE. Onychomycosis: Pathogenesis, diagnosis, and man-
agement. Clin Microbiol Rev 1998;11:415–429.
10. Elewski BE, Leyden J, Rinaldi MG, Atillasoy E. Office practice-
based confirmation of onychomycosis: A US nationwide prospect-
ive survey. Arch Intern Med 2002;162:2133–2138.
11. Nenoff P, Ginter-Hanselmayer G, Tietz HJ. Onychomykose – ein
Update: Teil 2 – Vom Erreger zur Diagnose – konventionelle und
molekularbiologische mykologische Diagnostik. [Fungal nail infec-
tions – An update: Part 2: From the causative agent to diagnosis –
Conventional and molecular procedures]. Hautarzt 2012;63:
130–138.
12. Finch JJ, Warshaw EM. Toenail onychomycosis: Current and future
treatment options. Dermatol Ther 2007;20:31–46.
13. Epstein E. How often does oral treatment of toenail onychomycosis
produce a disease-free nail? An analysis of published data. Arch
Dermatol 1998;134:1551–1554.
14. Gupta AK, Ryder JE, Johnson AM. Cumulative meta-analysis of
systemic antifungal agents for the treatment of onychomycosis. Br J
Dermatol 2004;150:537–544.
15. Van Duyn GL, Elewski BE. Recent updates in oral terbinafine: Its
use in onychomycosis and tinea capitis in the US. Mycoses 2011;
54:e679–e685.
16. Tosti A, Piraccini BM, Stinchi C, Colombo MD. Relapses
of onychomycosis after successful treatment with systemic
Int J Hyperthermia, 2013; 29(6): 544–550
antifungals: A three-year follow-up. Dermatology 1998;197:
162–166.
17. Tchernev G, Penev PK, Nenoff P, Zisova LG, Cardoso JC, Taneva
T, et al. Onychomycosis: Modern diagnostic and treatment
approaches. Wien Med Wochenschr 2013;163:1–12.
18. Singal A, Khanna D. Onychomycosis: Diagnosis and management.
Indian J Dermatol Venereol Leprol 2011;77:659–672.
19. Grover C, Bansal S, Nanda S, Reddy BS, Kumar V. Combination of
surgical avulsion and topical therapy for single nail onychomycosis:
A randomized controlled trial. Br J Dermatol 2007;157:364–368.
20. Silva JL, Doimo G, Faria DP. The use of high frequency waves to
treat onychomycosis: Preliminary communication of three cases.
An Bras Dermatol 2011;86:598–600.
21. Aspiroz C, Fortuno CB, Rezusta A, Paz-Cristobal P, Dominguez-
Luzon F, Gene DJ, et al. Terapia fotodinámica aplicada al
tratamiento de las onicomicosis. Presentación de un caso y revisión
de la literatura. [Photodynamic therapy for onychomycosis. Case
report and review of the literature]. Rev Iberoam Micol 2011;28:
191–193.
22. Kamp H, Tietz HJ, Lutz M, Piazena H, Sowyrda P, Lademann J,
et al. Antifungal effect of 5-aminolevulinic acid PDT in
Trichophyton rubrum. Mycoses 2005;48:101–107.
23. Rothermel E, Apfelberg DB. Carbon dioxide laser use for certain
diseases of the toenails. Clin Podiatr Med Surg 1987;4:809–821.
24. Kaplan I, Labandter H. Onychogryphosis treated with the CO2
surgical laser. Br J Plast Surg 1976;29:102–103.
25. Apfelberg DB, Rothermel E, Widtfeldt A, Maser MR, Lash H.
Preliminary report on use of carbon dioxide laser in podiatry. J Am
Podiatry Assoc 1984;74:509–513.
26. Vural E, Winfield HL, Shingleton AW, Horn TD, Shafirstein G.
The effects of laser irradiation on Trichophyton rubrum growth.
Lasers Med Sci 2008;23:349–353.
27. Kimura U, Takeuchi K, Kinoshita A, Takamori K, Hiruma M,
Suga Y. Treating onychomycoses of the toenail: Clinical efficacy of
the sub-millisecond 1064 nm Nd: YAG laser using a 5 mm spot
diameter. J Drugs Dermatol 2012;11:496–504.
28. Hiruma M, Kawada A, Noguchi H, Ishibashi A, Conti Diaz IA.
Hyperthermic treatment of sporotrichosis: Experimental use of
infrared and far infrared rays. Mycoses 1992;35:293–299.
29. Bornstein E, Hermans W, Gridley S, Manni J. Near-infrared
photoinactivation of bacteria and fungi at physiologic temperatures.
Photochem Photobiol 2009;85:1364–1374.
30. Landsman AS, Robbins AH, Angelini PF, Wu CC, Cook J,
Oster M, et al. Treatment of mild, moderate, and severe
onychomycosis using 870- and 930-nm light exposure. J Am
Podiatr Med Assoc 2010;100:166–177.
31. Manevitch Z, Lev D, Hochberg M, Palhan M, Lewis A, Enk CD.
Direct antifungal effect of femtosecond laser on Trichophyton
rubrum onychomycosis. Photochem Photobiol 2010;86:476–479.
32. Hochman LG. Laser treatment of onychomycosis using a novel
0.65-millisecond pulsed Nd:YAG 1064-nm laser. J Cosmet Laser
Ther 2011;13:2–5.
33. Kozarev J, Vizintin Z. Novel laser therapy in treatment of
onychomycosis. J Laser Health Acad 2010;1:1–8.
34. Hees H, Raulin C, Baumler W. Laser treatment of onychomycosis:
An in vitro pilot study. J Dtsch Dermatol Ges 2012;10:913–918.
35. Tietz HJ, Nenoff P. Die Onychomykose – ein Kronjuwel der
Dermatologie. [Onychomycosis: A crown jewel of dermatology].
Hautarzt 2012;63:842–847.
36. Kozarev J. ClearSteps – Laser onychomycosis treatment:
Assessment of efficacy 12 months after treatment and beyond.
J Laser Health Acad 2012;2011:S07.