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Journal of Clinical Microbiology, March 2000, p. 1263-1266, Vol. 38, No. 3
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Otitis Externa Associated with Malassezia
sympodialis in Two Cats
M. J.
Crespo,
M. L.
Abarca, and
F. J.
Cabañes*
Departament de Patologia i Producció
Animals (Microbiologia), Facultat de Veterinària, Universitat
Autònoma de Barcelona, 08193-Bellaterra (Barcelona), Spain
Received 23 June 1999/Returned for modification 19 November
1999/Accepted 20 December 1999
 |
ABSTRACT |
The lipid-dependent species Malassezia sympodialis was
isolated from two cats with otitis externa. To our knowledge, this is
the first report of the isolation of lipid-dependent species of the
genus Malassezia associated with skin disease in domestic animals.
| |
TEXT |
Malassezia species are
lipophilic yeasts that form an integral part of animal and human
cutaneous microbiota (19). They are also considered
important medical yeasts because they are etiological agents of chronic
skin disorders and have been reported with increasing frequency as
causative agents of life-threatening, iatrogenic, catheter-related
sepsis both in immunocompromised individuals (1) and
low-birth-weight neonates receiving parenteral lipid alimentation
(23).
The genus has recently been revised on the basis of molecular data and
lipid requirements and enlarged to include seven species. They include
the three former species Malassezia furfur (Robin) Baillon
1889, Malassezia pachydermatis (Weidman) Dodge 1935, and Malassezia sympodialis Simmons & Guého 1990 and four
new taxa, Malassezia globosa, Malassezia obtusa,
Malassezia restricta, and Malassezia slooffiae
(9, 12). M. pachydermatis is the only species not
dependent on lipid supplementation for in vitro growth; the other six
species are lipid dependent.
All lipid-dependent species can be isolated from healthy and diseased
human skin. They may be involved in skin disorders such as pityriasis
versicolor, folliculitis, seborrhoeic dermatitis, and atopic dermatitis
and even in systemic infection (8). M. pachydermatis is often isolated from domestic and wild animals (11), and it has occasionally been implicated in cases of
systemic infection in humans (18, 24). This species can play
an important role in chronic dermatitis and otitis externa, especially
in carnivores. It is the most common yeast that contributes to otitis
externa as a perpetuating factor in dogs and cats (20).
However, otitis externa associated with lipid-dependent species in cats
or other animals has not been mentioned to date.
In this paper, otitis externa associated with the lipid-dependent
species M. sympodialis in two cats is reported. To our
knowledge, this is the first report of the isolation of M. sympodialis involved in a skin disorder in domestic animals.
Case 1.
A 10-year-old female Persian cat presented with acute
otitis externa in the right ear. The animal had pruritus and an
excessive aural discharge. On otoscopic examination, a generalized
erythema was seen, and the external meatus was full of flaky black wax. On examination, the cat did not have any other dermatological disorders
or pathologies. The animal had no history of skin diseases.
Case 2.
A 14-year-old female Angora cat presented with a
prolonged period of heat with weight loss and chronic otitis externa in
the left ear with pruritus. The external ear canal was full of brownish wax, and erythema was seen on otoscopic examination. No other dermatological disorders or pathologies were detected. The cat did not
have a history of skin diseases.
Microbiology.
Swabs from the external ear canals of the two
cats were obtained for microbiologic examination. The smears were
stained with Gram and Diff-Quick stains. In both cases, Gram and
Diff-Quick stains revealed the presence of numerous
Malassezia cells, more than 10 organisms per high-power
field. Hyphae were not seen. Buds were formed on a narrow base (Fig.
1), which differed from the monopolar
budding on a broad base typical of M. pachydermatis (Fig.
2). No bacteria were detected in any
case. Cultures on Sabouraud glucose agar (SGA), SGA supplemented with
olive oil (10 ml/liter), and Leeming's medium (10 g of peptone, 5 g of glucose, 0.1 g of yeast extract, 4 g of desiccated ox
bile, 1 ml of glycerol, 0.5 g of glycerol monostearate, 0.5 ml of
Tween 60, 10 ml of whole-fat cow's milk, 12 g of agar per liter
[pH 6.2]) (14) were made for mycological examination. All
media contained 0.05% chloramphenicol and 0.05% cycloheximide. In the
first case, the sample was inoculated only on these media because a
presumed otitis due to M. pachydermatis had been clinically
diagnosed. In the second case, the sample was also inoculated on blood
agar and MacConkey agar for bacteriological examination. Plates of SGA,
SGA supplemented with olive oil, and Leeming's medium were incubated
at 35°C and examined after 3, 5, 7, and 14 days. Plates of blood agar
and MacConkey agar were incubated at 37°C with 5% CO2
for 3 days.
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FIG. 1.
Gram stain of a smear from an otic swab showing the
presence of numerous M. sympodialis cells. Note that buds
are formed on a narrow base. Bar, 10 µm.
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FIG. 2.
Diff-Quick stain of a smear from an otic swab showing
the typical monopolar budding on a broad base of M. pachydermatis. Bar, 10 µm.
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Cultures on SGA supplemented with olive oil and Leeming's medium
yielded numerous yeast colonies at 5 days of incubation. Two different
types of colonies were isolated on SGA supplemented with olive oil: one
type was yellow with a creamy texture, and the other was white with an
oily texture. On Leeming's medium, all colonies were white with a soft
texture. Cultures on SGA were negative at 14 days of incubation.
Bacteriological culture was negative at 3 days of incubation (case 2).
Five different colonies were selected from SGA supplemented with olive
oil (including the two different types of colonies) and Leeming's
medium and subcultured onto SGA to determine their lipid dependence.
All selected yeasts were considered lipid dependent species because
they failed to grow on SGA. The identification of these lipid-dependent
yeasts was based on the ability to use certain polyoxyethylene sorbitan
esters (Tweens 20, 40, 60, and 80), following the key for
identification of species described by Guého et al.
(9) and the Tween diffusion test proposed by Guillot et al.
(13). The Cremophor EL assimilation test and the splitting
of esculin described by Mayser et al. (16) were used as
additional key characters for the differentiation of the species
M. furfur, M. slooffiae and M. sympodialis. All isolates formed cream-colored, smooth, and
glistening colonies with average diameters of 1.2 to 2.6 mm on modified
Dixon's agar (36 g of malt extract, 6 g of peptone, 20 g of
desiccated ox bile, 10 ml of Tween 40, 2 ml of glycerol, 2 ml of oleic
acid, 12 g of agar per liter [pH 6.0]) (9) after 7 days of incubation at 32°C. These data differed from the 5-mm average
diameter described by Guého et al. for this species under
the same conditions (9). The cells were ovoid to globose
(1.4 to 1.9 µm by 2.0 to 2.7 µm), and buds were formed on a
narrow base. Sympodial budding was seen in some cells. They exhibited a
positive catalase reaction. In the Tween diffusion test, all isolated
yeasts utilized the four Tweens (20, 40, 60, and 80) showing inhibition
areas around the Tween 80 and Tween 20 wells, as did the type strain
M. sympodialis CBS 7222T under the same
conditions. The isolates grew on glucose-peptone agar with 0.5% Tween
60 and 0.1% Tween 80, and they did not grow on 10% Tween 20. They did
not assimilate Cremophor EL, and the splitting of esculin was strongly
positive in 2 to 3 days. According to these findings, all
lipid-dependent yeasts were identified as M. sympodialis
(9, 13, 16).
Susceptibility tests on one isolate of each case were performed with
antifungal tablets (Neo-Sensitabs; Rosco Diagnostica, Taastrup,
Denmark) (5), using modified Dixon's agar. Inocula containing about 109 cells ml
1 were prepared
from a fresh culture on modified Dixon's agar with 5 days of
incubation at 32°C. The strains were sensitive to amphotericin B,
ketoconazole, fluconazole, itraconazole, miconazole, and clotrimazole. All isolates were resistant to 5-fluorocytosine.
Treatment.
The cats were treated topically with Conofite Forte
(Esteve Veterinaria, Laboratorios Dr. Esteve S. A., Barcelona,
Spain) (20 mg of miconazole, 5,000 IU of polymyxin B, 5 mg of
prednisolone [each per ml]), 3 to 5 drops/ear twice a day for 10 days. The otitis successfully resolved after treatment of both cats.
Discussion.
Among the different Malassezia species,
the non-lipid-dependent species M. pachydermatis can be
isolated from the external ear canal and mucosae of healthy cats and
cats with otitis externa and dermatitis (11). This species
is considered a nonpathogenic, normal commensal organism which can
become an opportunistic pathogen when the microclimatic factors are
appropriate or the host's defense mechanism fails or is overwhelmed
(15, 17). However, it is more frequently isolated from dogs
than cats. In cats, the relative importance of M. pachydermatis in disease is less certain because it is found with
equal frequency in clinically healthy cats and in those with otitis
externa. M. pachydermatis has been identified in 50 to 83%
of dogs with otitis externa and in 19% of cats with otitis externa
(7).
In addition to this non-lipid-dependent species, the lipid-dependent
species M. sympodialis (3), M. globosa
(4), and M. furfur (6) may also
colonize the skin and mucosae of healthy cats. In fact, cats and
cheetah are the only carnivores in which the isolation of
lipid-dependent species had been demonstrated (10).
Recently, lipid-dependent species have been also reported in mixed
cultures from canine and feline specimens, using identification techniques such as the Tween 80 diffusion test, the Cremophor EL
assimilation test, and the splitting of esculin (22). In our
opinion, pure cultures must be used to obtain a microbiological identification following this kind of technique. On the other hand, the
role of lipid-dependent species in animal skin is still unclear. It has
been mentioned that they can colonize the skin of different animals,
but no skin disorder associated with lipid-dependent species in
domestic animals has been mentioned to date, with the exception of a
skin disorder in goats with the same typical lesions as tinea
versicolor in humans (2). However, in that case, the isolation in vitro of the causal agent was unsuccessful. To our knowledge, our description herein is the first report of the isolation of lipid-dependent species of the genus Malassezia
associated with skin disease in domestic animals. To date, this is the
first isolation of M. sympodialis involved in otitis externa
in cats.
The two cases of otitis were resolved successfully by topically
treating the cats with Conofite Forte, which includes miconazole among
other drugs. As previously mentioned, the isolates were sensitive to
miconazole. Although in general, the in vitro antifungal susceptibilities of the different pathogenic fungi can be a valuable guide for the practitioner, reliable antifungal susceptibility testing
is still poorly developed, especially for lipophilic yeasts. Recently,
some testing conditions have been proposed as guidelines for a
reference broth microdilution method (21), but the yeasts of
the genus Malassezia are not included.
M. sympodialis seems to be the most lipid-dependent species
isolated from healthy cats (3). Although M. pachydermatis can be involved in otitis externa in cats, it does
not occur with the frequency it does in cases of otitis externa in
dogs. Usually, practitioners diagnose this kind of otitis in the
laboratory by microscopic examination of stained smears from otic
swabs. For this reason, the presence of lipid-dependent
Malassezia species in the smears may be ignored and/or the
organism may be misidentified as M. pachydermatis.
Nevertheless, as has been pointed out, this species has a different
micromorphology. On the other hand, since culture media with lipid
sources, such as SGA supplemented with olive oil, Leeming's medium,
and modified Dixon's agar, are often recommended for the isolation of
M. pachydermatis (7), the isolated yeasts could
be misidentified as M. pachydermatis because their lipid
dependence is not tested in a routine way. All of the arguments
described above allow us to think that M. sympodialis could
be more common in this microenvironment and could play a similar role
as M. pachydermatis in otitis externa in cats.
| |
ACKNOWLEDGMENTS |
We thank Ana Avellaneda (Ars Veterinaria) and Javier Mora
(Consultori Veterinari Horta) for the samples kindly provided for this investigation.
| |
FOOTNOTES |
*
Corresponding author. Mailing address: Departament de
Patologia i Producció Animals (Microbiologia), Facultat de
Veterinària, Universitat Autònoma de Barcelona,
08193-Bellaterra (Barcelona), Spain. Phone: 34 93 581 10 88. Fax: 34 93 581 20 06. E-mail: javier.cabanes{at}uab.es.
| |
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Journal of Clinical Microbiology, March 2000, p. 1263-1266, Vol. 38, No. 3
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
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