CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASE
Original Article
Clinical Comparison of the Baxter MicroScan
Yeast Identification Panel® and the Vitek
Yeast Biochemical Card®
DONNA L. RIDDLE, MS, MT(ASCP), 1 OLARAE GIGER, P H D , 2 LINDA MILLER, P H D , 3
GERRI S. HALL, P H D , 4 AND GAIL L. WOODS, MD 5
croScan panels, 24-48 hours with Vitek cards, and 72 hours with the
To determine the reliability of the Baxter MicroScan Yeast IdentificaAPI 20C strips. On initial testing, 101 (67%) and 128 (85%) isolates,
tion Panel,™ processed by the Walkaway-96, and the Vitek Yeast Biorespectively, were correctly identified by MicroScan and Vitek. After
chemical Card,"* 150 clinical yeast isolates (30 Candida albicans, 67
Candida species, not albicans, 26 Torulopsis glabrata, 13 Cryptococcus repeat testing, the number of correctly identified isolates increased to
neoformans, 4 Saccharomyces cerevisiae, 6 Trichosporon beigelii, 3123 (82%) by MicroScan and to 142 (95%) by Vitek. Yeasts most comRhodotorula species, and 1 Geotrichum species) were tested on both monly misidentified were Candida tropicalis, Tglabrata, and Candida
systems. Results were compared with those obtained by the API 200""
parapsilosis by MicroScan and C tropicalis and T glabrata by Vitek.
(Key words: Automation; Rapid microbiology; Yeast identification) Am
and the appearance of yeast cells on cornmeal Tween-80 agar. After
J Clin Pathol 1994;101:438-442.
inoculation of each system, results were available in 4 hours with Mi-
groups, have been developed. Growth-based systems that rely
primarily on the ability of the organism to assimilate various
substrates include the API 20C Clinical Yeast System (Analytab Products, Plainview, NY) and the Vitek Yeast Biochemical
Card (Vitek Systems, Hazelwood, MO).7"12 Other systems,
such as the Baxter MicroScan Yeast Identification Panel
(Baxter MicroScan, West Sacramento, CA), use chromogenic
and modified conventional tests to assess the presence of preformed enzymes.13"15
The purpose of this study was to evaluate the reliability of
two commercially available automated systems: The Baxter
MicroScan Rapid Yeast Identification Panel, processed by the
Walkaway-96, and the Vitek Yeast Biochemical Card. The
identification given by each system was compared with the
reference method, the API 20C Clinical Yeast System, and the
morphologic appearance of yeast cells on cornmeal Tween-80
(CMT-80) agar.
Over the last several years, yeasts have become recognized as
significant opportunistic pathogens. The need for efficient, reliable methods for identification of yeasts has therefore become
increasingly important. 1 " 3 Factors that compromise the normal
host defenses increase the risk of serious yeast infection. These
factors include use of cytotoxic drugs, steroids, or other immunosuppressive agents; infection with human immunodeficiency virus; intravenous drug use; placement of intravascular
catheters; and intra-abdominal surgery.
Because the susceptibility of yeasts to antifungal agents varies, appropriate therapy for invasive yeast infections may depend on the rapid and accurate identification of the responsible
organism. In vitro, for example, Candida lusitaniae is resistant
to amphotericin B, and Candida krusei and Torulopsis glabrata are resistant to fluconazole.4"6
Classic methods of yeast identification are technically complex and time consuming, often requiring incubation for up to
1 week. The result, therefore, may be of retrospective interest
only. Over the last 2 decades, commercial systems for more
rapid yeast identification, which can be divided into two basic
MATERIALS A N D METHODS
Organisms
One hundred fifty clinical yeast isolates were tested; 98 from
From the'Department of Pathology and Laboratory Medicine, Hos- cultures at the Cleveland Clinic Foundation, Cleveland, Ohio,
2
pital ofthe University of Pennsylvania. Philadelphia. Pennsylvania; De- and 52 from the Medical College of Pennsylvania, Philadelpartment of Pathology and Laboratory Medicine. Episcopal Hospital,
Philadelphia. Pennsylvania; ^Department ofPathology and Laboratory phia, Pennsylvania. Each isolate was subcultured to Sabouraud
Medicine. Holy Redeemer Hospital and Medical Center, Philadelphia, dextrose agar (SAB) at least twice before it was tested by the
Pennsylvania; "Department ofMicrobiology, Cleveland Clinic Founda- three systems. MicroScan panels were evaluated at the Medical
tion, Cleveland, Ohio; and Department of Pathology and Laboratory College of Pennsylvania; Vitek cards, at the Cleveland Clinic
Medicine, Medical College of Pennsylvania, Philadelphia, Pennsyl- Foundation; and the API 20C strips and CMT-80 agar, at both
vania.
institutions.
Manuscript received November 30, 1992; revision accepted March
8, 1993.
Address reprint requests to Dr. Woods: Department of Pathology,
University of Texas Medical Branch, Galveston, TX 77555-0743.
MicroScan Rapid Yeast Identification
Panel
Isolates were subcultured to SAB and incubated 48-72 hours
at 25-30 °C in ambient air. Testing was performed according
438
RIDDLE ET AL.
Baxter MicroScan Versus •k Yeast Biochemical Card
to manufacturer's directions. Isolated colonies of yeast were
inoculated into 3 mL MicroScan Inoculum Water to give a
turbidity approximately equal to a number 8 McFarland barium sulfate standard. The panels were labeled using preprinted
bar code labels generated by the Baxter MicroScan WalkAway
System. Using an Eppendorf (Brinkman Instruments, Inc.,
Westbury, NY) step pipettor, 50 jiL of the yeast suspension was
added to each of the 27 wells containing a substrate, the /3naphthylamide control well, the nitrophenyl control well, and
the locator well. Each panel was then loaded into the WalkAway, which incubated panels at 35 °C, for processing.
A base-line reading of each panel was performed after 30
minutes. After 4 hours, the panel was again drawn into the
instrument read head, where reagents were added and final
reactions interpreted. Positive or negative reactions were based
on the color change in each well, compared with the base-line
reading. As recommended by the manufacturer, no visual verification of the WalkAway interpretations or editing of panel
results was attempted. The nine-digit biotype number generated by assigning a weighted numerical value to all positive
reactions was compared with biotype numbers included in the
computer database of the instrument. Based on this comparison, the computer provided an identification and a percent
probability of its accuracy. According to manufacturer's guidelines, an identification with a less than 85% likelihood of being
correct is questionable.
Vitek Yeast Biochemical Card
Vitek Yeast Biochemical Cards were tested according to the
manufacturer's directions. Isolates were grown at 28-30 °C on
SAB for 18-48 hours, or until sufficient growth was achieved.
Several well-isolated, identical colonies were picked and emulsified into 1.8 mL of .45% to .5% saline to give a turbidity equal
to a number 2 McFarland standard. Cards werefilledwith the
yeast suspension using the filling module of the Vitek instrument, incubated off-line at 30 °C for 24 hours, and then loaded
into the Vitek reader/incubator for one reading. If additional
incubation was required for identification, cards were reincubated and read again at 48 hours. A well was called positive or
negative based on the amount of light transmitted through the
corresponding negative control well. The nine-digit biotype
number, generated by assigning a weighted numerical value to
all positive reactions, was compared with those included in the
computer database, and an identification was determined and
given a probability of being correct.
API 20C Clinical Yeast System
The API 20C was performed according to the manufacturer's directions. Isolates were grown on SAB for 48-72 hours
at 25-30 °C. Well-isolated colonies were emulsified in an ampule of melted API 20C basal medium to give density just below a 1+ on a Wickerham card. Before the basal medium
cooled to a solidified state, each well of the API 20C strip was
inoculated, and the strip was placed in an incubation tray, covered with a lid, and incubated at 30 °C for 72 hours. Reactions
were visually examined at 24,48, and 72 hours and recorded as
positive or negative, based on the turbidity in the well. A sevendigit biotype number was generated by assigning a weighted
score to positive reactions. Identification was based on the gen-
439
erated biotype code, which was compared with those listed in
the API 20C Analytical Profile Index.
Cornmeal Tween-80
The microscopic appearance of yeast cells on CMT-80 agar
was considered in the identification of each isolate. With a
sterile inoculating needle, yeast cells were picked from isolated
colonies and inoculated to CMT-80 agar by using the needle to
cut two to three parallel lines no less than 1 cm apart into the
agar. A portion of the cuts was covered with a coverslip. Plates
were incubated at 25-30 °C for 18-48 hours and then examined at X 100 to X 400 for characteristic structures (blastoconidia, arthroconidia, chlamydospores, ascospores, true hyphae,
and pseudohyphae).16
Resolution of Discrepancies
In this study, the identification provided by the MicroScan
panel or the Vitek card was considered correct if the following
criteria were fulfilled: genus and species agreed with those provided by the API 20C, structures observed on CMT-80 agar
were consistent with the identification, and the likelihood that
the identification was correct was 85% or greater. The panel or
card was repeated using a fresh subculture if either automated
system did not provide an identification, if the probability that
the identification was correct was less than 85%, if the identification did not correlate with the structures observed on CMT80 agar, or if additional off-line conventional testing other than
CMT-80 agar was required for confirmation. If, after repeat
testing, the genus and species provided by the MicroScan or
Vitek still did not fulfill the above criteria for correct identification, the result was considered incorrect.
RESULTS
On initial testing, 101 (67%) and 128 (85%) isolates, respectively, were correctly identified by MicroScan and Vitek. After
repeat testing (based on the criteria previously described), the
number of correctly identified isolates increased to 123 (82%)
by MicroScan and 142 (95%) by Vitek. The results are summarized in Tables 1 and 2, respectively.
The time required for final identification varied for each system evaluated. All three systems recommend subculture from
primary isolation media to SAB to obtain pure growth of the
yeast to be tested. The suggested incubation period for that
subculture is 18-72 hours for MicroScan (depending on the
growth of the isolate), 18-48 hours for Vitek, and 48-72 hours
for the API 20C. After inoculation of the system, the time to
identification is 4 hours for MicroScan, 24-48 hours for Vitek,
and 72 hours for the API 20C (an isolate may be identified by
the API 20C in 24-48 hours, but the manufacturer's guidelines
require incubation for 72 hours). Total time to identification,
therefore, is 22-76 hours for MicroScan, 42-96 hours for Vitek, and 72-144 hours for the API 20C.
DISCUSSION
Commercial systems for identification of yeasts are technically simple to perform and provide a much shorter turn-
Vol. 101 •No. 4
440
CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASE
Original Article
TABLE 1. ACCURACY OF THE MICROSCAN RAPID YEAST IDENTIFICATION PANEL COMPARED WITH THE API 20C
CLINICAL YEAST SYSTEM AND CORNMEAL TWEEN 80 AGARt (CMT-80)
API 20C/CMT-80
Identification
Candida spp.
C albicans
C. guilliermondii
C. krusei
C. lusitaniae
C parapsilosis
C. paratropicalis
C. pseudotropicalis
C. rugosa
C. tropicalis
Cryptococcus neoformans
Geotrichum spp.
Rhodolorula spp.
Saccharomyces cerevisiae
Torulopsis glabrata
Trichosporon beigelii
Total
Total
Tested
No. (%) Correctly
Identified on
Initial Testing
30
2
5
2
19
2
1
1
35
13
1
3
4
26
6
30(100)
1(50)
1(20)
0(0)
12(63)
2(100)
1 (100)
0(0)
17(49)
12(92)
1 (100)
0(0)
4(100)
16 (62)
4(67)
101 (67)
150
around time than conventional methods. They should not be
used, however, if results are not accurate. We therefore evaluated the reliability of two commercially available automated
yeast identification systems: MicroScan and Vitek. In our
study, the accuracy of these instruments was determined by
testing clinical isolates (15 different yeasts) and comparing the
results obtained by each instrument to the results of a reference
method: the API 20C Clinical Yeast System and the appearance of yeast cells on CMT-80 agar, which has been used as the
reference method in several clinical evaluations of commercial
yeast identification systems."'12'1415 Our goal was to assess the
performance of the MicroScan and Vitek as stand-alone systems in a clinical laboratory; therefore, rarely encountered
No. Correctly
Identified on
Repeat Testing
—
1
2
1
3
—
—
0
7
1
—
1
—
6
1
22
Total (%)
Correctly
Identified
30(100)
2(100)
3(60)
1(50)
15(79)
0(0)
1 (100)
0(0)
24 (69)
13(100)
1 (100)
1(33)
4(100)
22 (85)
5(83)
123(82)
yeasts were not included, and no additional tests (other than
CMT-80 agar) were done.
On initial testing, the MicroScan WalkAway correctly identified only 67% of the 150 isolates in the study, which is considerably fewer than the number found by other investigators who
evaluated the MicroScan panels.1415 In a study conducted by
Land and colleagues,14 MicroScan panels correctly identified
92% of those organisms included in the database and 85% of all
yeasts tested. Of the yeasts that these investigators categorized
as uncommon or "not in database" (and, therefore, excluded
from the calculation of 92% overall agreement), only Candida
rugosa (1 isolate) and Rhodolorula species (3 isolates) were
included in our study. All four were misidentified on initial
TABLE 2. ACCURACY OF THE VITEK YEAST BIOCHEMICAL CARD COMPARED WITH THE API 20C CLINICAL YEAST
SYSTEM AND CORNMEAL TWEEN 80 AGAR (CMT-80)
API 20C/CMT-80
Identification
Candida spp.
C. albicans
C. guilliermondii
C. krusei
C. lusitaniae
C. parapsilosis
C. paratropicalis
C. pseudotropicalis
C. rugosa
C. tropicalis
Cryptococcus neoformans
Geotrichum spp.
Rhodolorula spp.
Saccharomyces cerevisiae
Torulopsis glabrata
Trichosporon beigelii
Total
Total
Tested
30
2
5
2
19
2
1
1
35
13
1
3
4
26
6
150
No. (°fo) Correctly
Identified on
Initial Testing
No. Correctly
Identified on
Repeat Testing
30(100)
0(0)
3(60)
1(50)
18(95)
0(0)
1 (100)
0(0)
30 (86)
13(100)
1 (100)
3(100)
4(100)
20 (77)
3(50)
—
127 (85)
A.J.C.P.-April 1994
2
0
1
1
2
—
1
3
—
—
—
—
3
2
15
Total (%)
Correctly
Identified
30(100)
2(100)
3(60)
2(100)
19(100)
2(100)
1 (100)
1(100)
33 (94)
13(100)
1 (100)
3(100)
4(100)
23 (77)
5(83)
142 (95)
RIDDLE ET AL.
Baxter MicroScan Versus Vitek Yeast Biochemical Card
testing. If these four isolates were excluded from our study, the
accuracy of the MicroScan would increase from 67% (101/150)
to 69% (101/146) on initial testing and from 82% (123/150) to
84% (122/146) on repeat testing (one isolate of Rhodotorula
species was correctly identified on retesting).
After the study by Land et al,14 the MicroScan panel was
revised: isoleucine, urea, N-acetyl-galactosamine, and trehalose were reformulated for ease and consistency of reading.
This new panel was used in our study and in an evaluation by
St.-Germain and Beauchesne.15 The latter investigators reported an overall accuracy of the MicroScan panels (including
repeat testing) of 96.6%, compared with the API 20C plus additional tests: Wickerham broth carbohydrate and nitrate utilization, carbohydrate fermentation, phenol oxidase, urease, cycloheximide sensitivity, growth at 37-42 °C, germ-tube
formation, and morphologic appearance of yeast cells on
CMT-80 agar. Without these additional tests, however, only
78% of 357 isolates were identified correctly, similar to the 82%
accuracy we found on repeat testing. Our study was designed to
evaluate MicroScan panels as a stand-alone system, because a
primary advantage of these panels is the 4-hour turnaround
time, which would be markedly prolonged if additional conventional biochemical tests were required. The only test other than
MicroScan panels that we included was morphologic appearance of yeast cells on CMT-80 agar. CMT-80 agar was inoculated from growth on the primary isolation media (at the time
of subculture to SAB) and examined on the same day that the
MicroScan panels were inoculated and interpreted; thus, identification of the isolate was not delayed.
Three of the 15 different yeasts included in our study accounted for 53% (80/150) of isolates tested and more than 70%
(33/49) of misidentifications on initial testing of panels by the
WalkAway: Candida tropicalis (18/35 [51%] were misidentified), Torulopsis glabrata (10/26 [48%] were misidentified),
and Candida parapsilosis (7/19 [37%] were misidentified).
Fourteen of the 18 C tropicalis isolates that were incorrectly
identified or were called C tropicalis with a probability of less
than 85% would have been correctly called C tropicalis (probability, 99.9%) had either or both of two reactions—glycyl-Lproline 4-methyoxy-|8-naphthylamide (GLPR) and 3-indoxyl
phosphate (IDX)—been interpreted as positive rather than negative. Three of the 18 isolates were called C tropicalis with a
probability of 70% to 79%; all had positive GLPR and negative
IDX results. If the IDX reaction had been positive, these isolates would also have been identified as C tropicalis with a
probability of 99.9%. The remaining C tropicalis isolate was
called Candida paratropicalis, based on a negative sucrose reaction.
The IDX and GLPR reactions were also important in the
identification of C parapsilosis. Of the seven isolates of C parapsilosis misidentified by the MicroScan panels, three would
have been correctly identified with a probability of 94% had
either or both of the IDX and GLPR reactions been interpreted
as positive rather than negative. For two isolates, positive results for both the IDX and GLPR were necessary for an identification of C parapsilosis with greater than 85% probability.
For the remaining two misidentified isolates, the IDX, GLPR,
and L-tyrosine-|8-naphthylamide (TYR) reactions required a
positive rather than negative interpretation for identification of
C parapsilosis of any probability (97.2% for one; the other,
"low selectivity" C parapsilosis, Torulopsis inconspicua, Candida zeylanoides).
441
Of the 10 incorrectly identified isolates of T glabrata, six
would have been correctly called T glabrata (probability,
> 93%), if, in addition to the other reactions, the L-alanine
4-methoxy-/3-naphthylamide (ALA) reaction had been interpreted as positive rather than negative. Incorrect identification
of the other four isolates occurred because of more than one
atypical reaction.
Had the reactions IDX and GLPR for the 17 isolates of C
tropicalis and five Cparapsilosis and the ALA reaction for the
six T glabrata consistently been called positive, the overall
correlation between the MicroScan panels and the API 20C/
CMT-80 would have increased from 67% (101/150) to 86%
(129/150) on initial testing and from 82% (123/150) to 92%
(129/150) on repeat testing. Data from previous studies
showed similar inconsistencies with the IDX reaction.14,15
Problems with the IDX reaction and, to a lesser extent, with the
GLPR and ALA reactions also occurred with the quality control isolates in this study. For quality control organisms to give
expected reactions with these substrates, panels often had to be
repeated.
Difficulty with interpretation of chromogenic substrates has
been described previously.141517 An automated system, which
eliminates the subjectivity of a technologist's visual reading,
would be expected to provide more accurate interpretations of
the panels. That expectation, however, was not met in this
study. Panels were interpreted by the totally automated WalkAway, yet the results were less accurate than those reported by
investigators who used visual interpretation only14 or visual
plus instrument (AutoScan-4) interpretation.15
One possible source of inconsistency of the MicroScan panels is the requirement for a heavy inoculum (approximately
equal to the turbidity of a number 8 McFarland standard). A
suspension of that turbidity is difficult to compare accurately,
by visual examination, to the standard inoculum. According to
the manufacturer, over- or underinoculation of the panels can
cause variations in chromogenic results, but the range outside
the standard density required to cause these variations is not
clearly defined.
Vitek cards performed better than MicroScan panels in our
evaluation; 85% of yeasts were correctly identified on initial
testing and 95% on repeat testing. Our results are comparable
with the 97.2% accuracy of the Vitek (including repeated cards)
previously reported by El-Zaatari and colleagues.12 In the latter
study, as in ours, the only additional off-line test was morphologic appearance of yeast cells on CMT-80 agar, which should
be inoculated at the time of subculture from primary isolation
media to SAB, so that it can be examined when the card is
interpreted and thus not delay identification.
For the 15 different species tested in this study, no disproportionate misidentifications, as occurred with the MicroScan
panels, were seen with the Vitek cards. The most frequently
misidentified yeasts, C tropicalis (five isolates) and T glabrata
(six isolates), accounted for 47.8% (11/23) of all misidentifications and for 40.7% (61/150) of all isolates. More than one
atypical reaction accounted for the misidentification of all five
isolates of C tropicalis. Of the six isolates of T glabrata that
were misidentified on initial testing, three were called "7" glabrata or Geotrichum capitatum," with no distinction between
them. One was identified as T glabrata with a probability of
64%. For these four isolates, the glycerol well was interpreted as
positive. Had glycerol been called negative, all four would have
been identified as T glabrata at a probability of 99%. The re-
Vol. 101 - N o . 4
442
CLINICAL MICROBIOLOGY AND INFECTIOUS DISEASE
Article
maining misidentifications of T glabrata were due to more
than one atypical reaction. Had the identification of Tglabrata
been more accurate, the overall agreement between Vitek cards
and API 20C/CMT-80 would have increased from 85% (127/
150) to 87% (131/150) on initial testing and from 95% (142/
150) to 97% (145/150) on repeat testing.
Candida albicans comprised 20% (30/150) of the isolates in
our study. For rapid, cost-effective identification of C albicans
in a clinical microbiology laboratory, the germ-tube test typically would be performed; use of a MicroScan panel, Vitek
card, or the API 20C would be unnecessary, unless the isolate
was a germ tube-negative variant of C albicans. If isolates of C
albicans were excluded from our evaluation, the accuracy of
MicroScan panels would decrease to 59% (71/120) on initial
testing and to 77.5% (93/120) with repeat testing. The accuracy
of the Vitek card would decrease to 81% (97/120) on initial
testing and to 93% (112/120) on retesting.
In summary, each system that we evaluated has advantages
and disadvantages. The API 20C Clinical Yeast System is comparable with classic conventional techniques. It is a complete
system and requires no additional reagents or instrumentation.
Each strip costs approximately $5.60 (manufacturer's 1994 list
price). Quality control includes testing three different yeasts.
The API 20C, however, has the longest minimum turnaround
time, based on the manufacturer-recommended 72-hour incubation for final identification, and it is not automated. Reactions are interpreted by a technologist's visual examination of
turbidity only; therefore, observer subjectivity may affect consistency of results.
In our evaluation, the performance of the MicroScan system
was inferior to that of the Vitek system. The MicroScan system,
however, was the most rapid of those evaluated. Final identification of all yeasts included in the database is available 4 hours
after system inoculation. Panels may be interpreted either by
computer-based instrumentation or visually. To improve performance, several substrates on the MicroScan panel have been
reformulated since the kit was first available, but refinements
of additional substrates, such as 3-indoxyl phosphate, glycyl-Lproline 4-methoxy-/3-naphthylamide, and L-alanine 4-methoxy-/3-naphthylamide, should be considered to improve accuracy further. These panels are the most expensive of the
systems evaluated (1994 list price, $6.87), and quality control
requires testing seven organisms.
The Vitek Yeast Biochemical Card compared favorably with
the API 20C, and final identification is available 24 hours after
the card is inoculated for most common clinical yeast isolates.
Quality control requirements include testing four yeasts. Each
card costs approximately $4.60 (1994 list price). The major
disadvantage of the Vitek yeast identification system is the absolute requirement for instrumentation. Unless the laboratory
already uses a Vitek system for bacterial identification and sus-
ceptibility testing or is planning to do so, use of the Vitek cards
for yeast identification probably is not an option.
Acknowledgments. We thank Dawn Hare for her technical help.
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