Yeast colonization of mucous membranes 
Clinical study 
YEAST COLONIZATION OF MUCOUS 
MEMBRANES: 
PREV ALENCE AND IDENTIFICATION OF 
DIFFERENT SPECIES 
A. Stary, A. Kuchinka-Koch and A. Schiller 
ABSTRACT 
The prevalence of different Candida species on mucous membranes of the genital, perigenital, anal, or 
pharyngeal region was evaluated in 451 male and female patients. Furthermore, the reliability of CHROMagar™ 
Candida for the identification of certain yeast species, such as C. albicans, C. glabrata, C. tropicalis, C. krusei, 
and Saccharomyces cerevisiae was determined. The most prevalent yeast isolated from the genital tract was C. 
albicans (86.1%) followed by C. glabrata (8.5%), C. parapsilosis (3.7%), and Saccharomyces cerevisiae (4.2%). 
The perigenital and anal regions were colonized most frequently with C. albicans (87.5%) only. In contrast 
to these samples a large variety of different Candida spp. was cultured from the pharyngeal region. Mixed 
yeast cultures mainly with C. albicans together with C. glabrata or S. cerevisiae were observed more frequently 
with pharyngeal (11.3%) than with vulvovaginal sam ples ( 4.6% ). Ali 408 presumptive C. albicans strains could 
be identified on CHROMagar™ Candida and were confirmed either by rice agar, germ-tube test, or API 
resulting in a sensitivity and specificity of 100%, respectively. The same performance characteristics could be 
observed for C. glabrata, C. krusei, and C. tropicalis . The comparison of different methods demonstrate that 
CHROMagar™ Candida permits a reliable and highly sensitive as well as specific differentiation of C. 
albicans, C. glabrata, C. tropicalis, and C. krusei from other Candida spp. by characteristic appearance of 
colonies within few days. 
KEYWORDS 
yeasts, prevalence, mucous membranes, CHROMagarTM Candida, genital tract 
INTRODUCTION 
In the past few years increasing rates of colonization 
of mucous membranes with different yeast species 
have been observed. This fact may be due to 
debilitating diseases such as AIDS, diabetes mellitus, 
and cancer, as well as to the increasing use of 
broad-spectrum antibiotics or corticosteriods (1,2). 
Furthermore, high-estrogen-content contraceptives, 
acta dennatovenerologica A.P.A. Vol 7, 98, No 3-4 
pregnancy, and tight-fitting clothes are reported risk 
factors especially for increased vagina! Candida 
colonization whereas sexual intercourse alone has 
no influence on the vagina! colonization rate with 
yeasts (3,4). 
Candida albicans is the most frequently iso~ated 
yeast associated with mucocutaneous colonization 
(5). This yeast is responsible for up to 90 per cent 
of recurrent vulvovaginal candidiasis (VVC) (3,6). 
121 
Yeast colonization of mucous membranes 
Table l. Mixed yeast cultures isolated from the Jemale genital tract listed according to yeasts found. 
+ 
12 9 
However, elevated isolation rates of other yeast 
species such as C. glabrata or C. tropicalis from 
genital mucous membranes have been reported (7, 
8). 
Primary as well as acquired resistance of certain 
yeast pathogens to antifungal drugs requires rapid 
and reliable identification of a broad spectrum of 
yeast species (9). Traditionally, methods for the 
identification of yeasts rely on a combination of 
morphological and biochemical characteristics. Pota-
ssium hydroxide microscopy is a rapid test for 
diagnosis of yeasts in different specimen types with 
a sensitivity of approximately 70% but the Jack of 
identification poses limits (10). A rapid and cost-
effective technique for the identification of C. albicans 
is the germ-tube test which has been reported to be 
negative in up to 5% of C. albicans isolates (11, 
12). Since the differentiation of other Candida spp. 
is not possible by the germ-tube test, cultivation is 
necessary for further identification. There are different 
media available for the detection of chlamydospores 
and the production of hyphae or pseudohyphae 
permitting the identification of diverse Candida spp. 
as well as the differentiation of the genera Crypto-
coccus, Saccharomyces, Geotrichum, and Trichosporon 
based on microscopic morphological features. The 
principal biochemical criteria for the identification 
of different species are assimilation of carbohydrates 
or nitrate, and fermentation of sugars. Since some 
of these methods are expensive and time-consuming, 
evaluations of newly developed, rapid, and accurate 
identification systems were performed (13). 
Recently, the potentially useful, rapid, and cost-
effective Microbial Identification System (MIS) for 
aerobic gram-positive, gram-negative bacterial species, 
and yeasts has been scrutinized for accuracy in 
identification of diverse yeast species with the result 
that it cannot be taken into consideration as alternative 
identification system for clinical microbiology labora-
tories at the moment (14, 15). 
For routine laboratory use exact and time-saving 
identification systems are of great importance. Recent 
122 
+ 
4 1 
reports described the chromogenic isolation medium 
CHROMagar™ Candida for the presumptive identifi-
cation of C. albicans, C. krusei, C. glabrata, and C. 
tropicalis based on species specific enzyme reactions 
(16, 17, 18, and 19). The results reveal CHROMagar™ 
Candida superior to Sabouraud glucose agar in its 
ability to detect mixed yeast cultures and to suppress 
bacterial growth. 
In the present study, the reliability of the chromo-
genic differential culture medium for the identification 
of different yeast species as well as for the detection 
of mixed yeast cultures isolated from clinical samples 
was determined. Furthermore, the prevalence of 
various Candida species on mucous membranes of 
the genital, perigenital, anal, and pharyngeal region 
was evaluated. 
PATIENTS AND METHODS 
During a period of two months 474 samples 
positive for yeasts were collected from 451 female 
and male patients attending the Outpatients' Centre 
for STD. Most of the specimens were collected 
from the genital mucous membranes, namely 283 
vulvovaginal specimens and 70 samples from the 
urethra and/or penis. Furthermore, 97 pharyngeal, 
11 anal, and 13 perigenital specimens were collected. 
Sampling was carried out with a sterile cotton-swab 
or wire-loop followed by immediate inoculation in 
Sabouraud broth (Oxoid, Unipath LTD., Hampshire, 
England) independent of specimen type. Yeasts were 
cultivated in Sabouraud broth at 37°C for two days 
before inoculating BBL® CHROMagar™ Candida 
(Becton Dickinson, Microbiology Europe, 38240 Meylan 
Cedex, France) and rice extract agar plates (Becton 
Dickinson, microbiology Systems, Cockeysville, MD 
21030 USA), respectively. Identification of different 
yeast species was performed by distinctive colour 
and colony characteristics on BBL® CHROMagar™ 
Candida after incubation for two days at 37°C. 
Confirmation of yeasts identified by BBL® CHROMa-
acta dennatovenerologica A.P.A. Vol 7, 98, No 3-4 
Yeast co/onization of mucous membranes 
gar™ Candida, was assessed by microscopy of rice 
extract agar plates incubated for two days at room 
temper"ature, germ-tube test, API20C AUX (Bio-
Merieux ™, Marcy l'Etoile, France), and Krusei-Color® 
Fumouze agglutination kit (Fumouze Diagnostics, 
92600 Asnieres, France). 
RESULTS 
PREV ALENCE OF YEASTS ON MUCO US 
MEMBRANES 
The prevalence of various yeast species isolated 
from different mucous membranes in a total of 451 
patients was 86.1 % for C. albicans, 7.6% for C. 
glabrata, 5.1 % for S. cerevisiae, 3.2% for C. parapsilosis, 
1.7% for C. krusei, 1.3% for C. tropicalis, and 0.6% 
for C. guillie,mondii. Twenty-six (5.5%) samples positive 
for yeasts could not be further identified and are 
given as Candida spp. 
The distribution of different yeasts cultivated from 
the genital tract according to gender is shown in 
Fig. l. Mixed yeast cultures were detected in 13 
( 4.6%) out of 283 vulvovaginal samples but not in 
specimens obtained from the male genital tract 
(Tab. 1.). 
C albicans was isolated from 11 out of 13 perigenital 
samples (84.6%) but was not present in the only 
mixed yeast culture obtained from the perigenital 
region containing C. krusei, S. cerevisiae, and Candida 
Table 2. Yeasts isolated from the pharyngeal region. 
acta dennatovenerologica A .P.A. Vol 7, 98, No 3-4 
spp. In 11 specimens collected from the anal region 
only C. albicans (90.9%) and C. krusei (9.1 %) were 
identified. 
A large variety of different Candida spp. was 
cultivated from the pharyngeal region with mixed 
yeast cultures observed in 11 (11.3%) out of 97 
samples (Tab. 2.). 
RESULTS OF BBL CHROMAGAR™ 
CANDIDA 
A total of 528 yeasts out of 474 positive cultures 
could be diagnosed with CHROMagar™ Candida. 
Ali 408 presumptive C. albicans strains could be 
identified on CHROMagar™ Candida and were 
confirmed either by microscopy of rice agar plates, 
germ-tube test, or API20C AUX resulting in a 
sensitivity and specificity of 100%, respectively (Tab. 
3.). A total of 36 C. glabrata, 6 C tropicalis, and 8 
C krusei isolates could be distinguished from other 
Candida spp. by the characteristics of their colonies 
on the chromogenic medium. Confirmation of these 
non-albicans strains was performed by morphological 
criteria on rice agar plates (ali 50 isolates), and 
additionally by biochemical characteristics in API20C 
AUX (5 C. glabrata, 6 C tropicalis isolates) as well 
as by agglutination test specific for C. krusei (8 C. 
krusei isolates) resulting also in a sensitivity and 
specificity of 100%, respectively. Species, such as C. 
parapsilosis, C guillie,mondii, and S. cerevisiae could 
123 
Yeast colonization of mucous membranes 
100 
89 
90 -
80 - 4, 3 
70 ' 
60 
50 
40 -
30 
20 
9,5 
10 . 
oi-;3 ~ 
~ . 
. ~ 
,9 . 
c.i 
12,9 
4;f] 7, 1 2;:n 1,1 0, 7 
• Women 
• Men 
0,3 0,3 
Fig. l. Frequencies of yeasts isolated from genital 
mucous membranes. The different yeast species are 
given in per cent far Jemale • and male • patients. 
be identified by microscopy and API20C AUX only, 
although these isolates formed colonies with chara-
cteristic surfaces creme to pink coloured on CHROMa-
gar™ Candida. 
DISCUSSION 
In this study 451 patients colonized with yeasts on 
different mucous membranes were included showing 
C. albicans (86.1 % ) as the most prevalent yeast 
followed by C. glabrata (7.6%) and S. cerevisiae 
(5.1 % ). Samples collected from the genital tract 
showed that both, men and women are most frequently 
colonized with C. albicans (86.1 % ) which is in 
concordance with earlier reports (6, 20). While in 
women C. glabrata and S. cerevisiae were isolated 
frequently, males showed colonization with C. para-
psilosis more often (Fig. 1). In 12.9% of positive 
specimens from the male genital tract the yeast 
species could not be further identified. This may be 
partly due to the fact that men are often pre-
treated with topical antimycotics before culture is 
obtained resulting in poor growth and diminished 
development of characteristic morphological features 
such as formation of chlamydospores. C. albicans is 
published to be the most prevalent yeast in men 
with balanitis but little is reported about the prevalence 
of other yeasts than C. albicans colonizing the male 
genital tract (20). Genital candidiasis in men is 
frequently associated with the presence of vaginal 
yeast colonization in a sexual partner (21). Since 
the prevalence of symptomatic and asymptomatic 
vulvovaginal candidiasis (VVC) has increased, epidemi-
ologic studies were carried out to investigate variation 
in the yeast flora. After ali, observations of changed 
occurrence of vaginal yeast pathogens are controversial 
(22,23,24,25). A possible change in the prevalence 
of yeast species may be associated with the type of 
antimycotic therapy as well as the selection of more 
resistant Candida species as a result of inappropriate 
or incomplete course of therapy (3,4) . However, 
information on the prevalence of diverse yeast species 
Fig. 2. Coloured colonics of different Candida specics on 
CHROMagar™ Candida incubated far 72 h at 37 C. 
Table 3. Comparison of different methods far the identification of Candida albicans. 
Number of isolates 
342 
30 
8 
1 
27 
S 408 
1 not done 
124 
CHROMagar Candida 
+ 
+ 
+ 
+ 
+ 
408 
Formation of chlamydospo.res Germ-tube test APl20C AUX 
+ nd 1 nd 
+ nd 
+ + 
+ 
nd + 
342 38 36 
acta dermatovenerologica A.P A. Vol 7, 98, No 3-4 
Yeast colonization of rnucous rnernbranes 
on different mucocutaneous membranes is rare. Thus 
additional studies involving a broad spectrum of 
specimen types are certainly of great importance 
and interest. 
Results of this study revealed CHROMagar™ 
Candida as a reliable differentiation medium for C. 
albicans and C. glabrata, the most frequently cultivated 
yeasts from mucous membranes (3,5,7). For the 
identification of C. albicans this medium showed a 
sensitivity as well as specificity superior to traditional 
methods such as the tests for the production of 
germ-tubes or chlamydoconidia techniques reported 
to give negative results with certain C. albicans 
strains (Tab. 3) (11,12,26) . Thus the particular value 
of CHROMagar™ Candida is obvious considering 
that confirmatory tests may no longer be indicated 
(16). Nevertheless, it has to be mentioned that 
clinical isolates of the less common yeast C. dublini-
ensis, a species closely related to C. albicans, have 
been misidentified as atypical strains of C. albicans 
with CHROMagar™ Candida (18,27). 
Beside C. albicans, the identification of the predo-
minant non-albicans Candida species C. glabrata is 
of importance for patients with recurrent VVC 
especially in immunosuppressed patients since it 
may cause clinical therapy failures (3,4,28,29) . In 
the present study identification of C. glabrata isolates 
with CHROMagarTM Candida was performed with 
the same accuracy as for C. albicans, C tropicalis, 
and C. krusei. These observations are in agreement 
with the findings of Pfaller and co-workers (19). 
Although the prevalence of C. tropicalis and C. 
krusei is rather low (0.6% and 1.3%, respectively), 
identification of these species is necessary for treatment 
management. Resistance to antimycotic drugs is seldom 
but occurs with strains of C. albicans in AIDS 
patients, C. glabrata, C krusei, and C. tropicalis 
rendering these yeast species as medically important 
(3,9,30) . 
A further advantage of CHROMagar™ Candida is 
the detection of mixed yeast cultures isolated from 
clinical samples on the basis of the distinctive and 
strongly differentiated colony colours of different 
yeast species growing. The results observed with 
mixed yeast cultures mainly containing C. albicans 
together with C. glabrata or S. cerevisiae in the 
present study are in accordance with earlier reports 
but could be detected more frequently with pharyngeal 
(11.3%) samples than with vulvovaginal specimens 
where 4.6% contained more than one yeast species 
(Tab. 1 and 2) (16,19). 
In sum, this report presents additional evidence 
for the usefulness of CHROMagarTM Candida for 
medica! mycological laboratories. We showed that 
the most prevalent as well as medically important 
yeast species can be reliably differentiated from 
other yeast species and that the detection of mixtures 
of yeasts isolated from clinical specimens is facilitated 
with CHROMagar™ Candida. 
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AUTHORS' ADRESSES 
Angelika Stary, MD, assoc. professor of dermatology, Outpatients' Centre for Venero-Dermatological 
Diseases, Franz Jonas Platz 8/2/3, A-1221 Vienna, Austria 
Angelika Kuchinka-Koch, MSc, molecular biologist, same address 
Angelika Schiller, medica! assistant, same address 
acta dennatovenerologica A.P.A. Vol 7, 98, No 3-4 127