Documente Academic
Documente Profesional
Documente Cultură
The authors evaluated the performance of the MycArray Yeast ID (Myconostica Ltd,
UK) assay in the identification of a total of 88 yeast isolates recovered in culture as
compared to that obtained through routine methods. The turn-around time for species
identification directly from cultures by the MycArray was 6 hours, much quicker than
classical methods and all yeasts were correctly identified. In two cases a double identification including Saccharomyces cerevisiae was noted, but it was not confirmed by culture. The results show that MycArray Yeast ID can be a potential tool for rapid detection
and identification of Candida species.
Keywords blood cultures, yeast, identification, microarray
Introduction
Nosocomial mycoses have become a growing phenomenon over the last decades, and in the USA they increased
more than 207% between the end of the 1970s and the
beginning of the new millennium [1]. This increase is the
result of the growing number of iatrogenic or pathological
immunocompromising conditions, infant prematurity,
solid and hematological cancer, abdominal surgery, presence of medical devices, and the use of broad spectrum
Received 17 August 2011; Received in final revised form 7 November
2011; Accepted 6 December 2011
Correspondence: Claudio Farina, UOC Microbiologia e Virologia AO
Ospedale San Carlo Borromeo, via Pio II no. 3 - 20153 Milano, Italy.
Tel: 39 02402 22456; Fax: 39 02402 22829; E-mail: farina.claudio@
sancarlo.mi.it
2012 ISHAM
antibiotic therapies [2,3]. In particular, candidemia incidence is 0.51.4/1,000 (in Italy 0.38/1,000) patients/day
and higher in Intensive Care Units at up to 2/1,000
patients/day [3,4].
The current gold standard for detection of candidemia
is culture of blood samples (blood culture). However, this
is not only a time-consuming method, but its sensitivity
for early detection of fungemia has been reported to be
as low as 50% [5]. In order to overcome the limitations
of blood cultures, various molecular approaches for the
identification of pathogenic fungi from blood have been
developed.
Molecular techniques are targeted to detect Candida
species in a short period of time, with a high sensitivity and
specificity. Several PCR tests have been devised, such as
nested PCR, multiplex PCR, Taq-man PCR, Light-Cycler
DOI: 10.3109/13693786.2011.648216
550
Farina et al.
PCR and fluorescent PCR. However, many of these methods are often limited by the number of species that can be
detected in a single assay [610].
The application of a DNA microarray technology may
allow the discrimination of a wide range of pathogens in a
single assay. The aim of the present study was to evaluate
the diagnostic performance of a new microarray method,
MycArray Yeast ID (Myconostica Ltd, Manchester, UK)
and to compare its results with those obtained by culture
considering the advantage of a rapid identification of isolates after their recovery from blood.
551
Species
Probe ID
Candida albicans
MA001
MA040
MA041
MA003
MA042
MA009
C. dubliniensis
C. famata
(Debaryomyces hansenii)
MA052
C. glabrata
MA011
MA047
MA048
MA010
C. guilliermondii
MA053
C. krusei
(Issatchenkia orientalis)
C. kefyr
(Kluyveromyces marxianus)
MA006
MA046
MA008
MA051
C. parapsilosis group
MA013
MA014
MA049
MA036
MA037
MA035
MA004
MA005
MA043
MA044
MA045
MA038
MA039
MA027
C. parapsilosis
C. metapsilosis
C. parapsilosis group
C. pelliculosa
C. rugosa
C. tropicalis
C. utilis
Cryptococcus neoformans
MA028
None outside
C. parapsilosis group
D. moranus (2);
D. robertsiae (3);
D. udenii (4)
None
None
None
C. athenensis (2);
C. smithsonii (2)
C. athensensis (2)
None
K. lactis (4)
K. lactis (3);
P. anomala (5);
S. cerevisiae (5)
None
None
None
None
None
None
None
None
None
Histoplasma capsulatum
MA032
None
Saccharomyces cerevisiae
MA033
MA034
(Continued)
552
Farina et al.
Table 1 (Continued)
Species
Probe ID
Rhodotorula mucilaginosa
MA029
R. mucilaginosa (cont.)
MA030
Internal control
MA031
MA015
MA016
MA017
MA018
None
N/A
N/A
Results
The microarray system yielded unequivocal identifications for each of the reference species (Table 2), which
were the same as those obtained by conventional methods. The layout of the chip allows for rapid and accurate
species identification (Fig. 1). Isolates of C. albicans (n
41) were identified by all the three oligonucleotidic
probes (MA001, MA040 and MA041), with the exception of six which were detected only by two probes
(MA040 and MA041). C. parapsilosis (n 10) was similarly identified by three probes (MA013, MA014 and
MA049) and one by two (MA013 and MA049), with
C. glabrata (n 10) detected by three probes (MA011,
MA047 and MA048), C. tropicalis (n 3) by four
(MA004, MA043, MA044 and MA045) but one strain by
two probes (MA043 and MA045) and finally the identification of C. guilliermondii, C. krusei and Cryptococcus
neoformans established by two probes (MA010 and
MA053; MA006 and MA046; MA027 and MA028,
2012 ISHAM, Medical Mycology, 50, 549555
553
Table 2 Evaluation of the DNA microarray with standard laboratory procedures (VITEK2 and ATB ID 32 C).
Organisma
Tested species
Positive probe
Candida albicans
41
C. parapsilosis
11
C. glabrata
C. tropicalis
C. tropicalis
C. lusitaniae
C. guilliermondii
C. krusei
C. neoformans
C. albicans C. glabrata
C. tropicalis C. glabrata
10
2
1
2
1
1
3
7
1
C. albicans
C. albicans
C. parapsilosis
C. parapsilosis
C. glabrata
C. tropicalis
C. tropicalis
Negative
C. guilliermondii
C. krusei
C. neoformans
C. albicans and C. glabrata
C. tropicalis and C. glabrata
aClinical
Fig. 1 MycArray Yeast ID results. Column 1 shows the micro array profile. Column 2 shows the PDF output with the graphical representation of results.
Signal strengths (Y-axis) are normalized against the biotin marker controls. The number of each probe is shown below the X-axis and the organism is
named above each column. In this example, all the control reactions were successful and C. parapsilosis was identified. Lines: (A) Cryptococcus
neoformans; (B) C. albicans.
554
Farina et al.
Discussion
Fungemia represents the 4th most common cause of bloodstream infections (BSI) in hospitalized patients in the USA
and the 7th in Europe [11,12]. It is related to longer hospitalizations and overall candidemia accounts for a 3575%
mortality rate [13]. It is well documented that its prognosis
is strictly related to the initiation of therapy, and that mortality increases more than 25% if there are delays between
the blood collection and the report of Candida recovery of
more than 48 hours [13].
The most common etiologic agent is C. albicans, followed by C. glabrata, C. parapsilosis, C. tropicalis and
C. krusei, with these five species accounting for more than
90% of all human cases.
Several other fungi can also cause fungemia including
Cryptococcus spp., S. cerevisiae, Histoplasma capsulatum,
Rhodotorula spp. and other Candida species. In Italy, many
studies suggest that C. parapsilosis is second to C. albicans
as the most common species recovered, being much more
frequent than all other Candida species [14].
Determining the etiology is important in the selection
of appropriate antifungal therapy because non-C. albicans
Candida species are more frequent and generally more
resistant to common therapies. For example, C. krusei is
fluconazole resistant and Rhodotorula spp., H. capsulatum
and Cryptococcus spp. are resistant to echinocandins [15].
The identification of the yeasts isolated from blood
cultures to the species level has become more and more
important to assure the best clinical management of
fungemia patients. In fact, the clinical outcome is related
to the early initiation of therapy, and the timely administration of antifungal drugs [1618].
Differences in virulence and in antifungal drug susceptibility of Candida spp. make identification and MIC
determination very important for clinical management
Acknowledgements
This study was supported by the financial support of
Myconostica Ltd, UK, (grant number for financial aid: 231
(2010.03.19).
Declaration of interest: The authors report no conflicts of
interest. The authors alone are responsible for the content
and writing of the paper.
References
1 Martin GS, Mannino DM, Eaton S. Moss M. The epidemiology of
sepsis in the United States from 1979 through 2000. N Engl J Med
2003; 348: 15461554.
2 De Pauw BE. Increasing fungal infections in the intensive care unit.
Surg Infect (Larchmt) 2006; 7 (Suppl 2): 9396.
3 Darouiche RO. Candida in the ICU. Clin Chest Med 2009; 30:
287293.
4 Tortorano AM, Biraghi E, Astolfi A, et al. European Confederation of
Medical Mycology (ECMM) prospective survey of candidaemia:
report from one Italian region. J Hosp Infect 2002; 51: 297304.
5 Ahmad S, Khan Z, Mustafa AS, Khan ZU. Seminested PCR for diagnosis of candidemia: comparison with culture, antigen detection, and
10
11
12
13
555