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Journal of Clinical Microbiology, December 1999, p. 4186-4188, Vol. 37, No. 12
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Identification of Hafnia alvei with the MicroScan
WalkAway System
L. A.
Rodríguez,1,*
J.
Vivas,1
C. S.
Gallardo,1
F.
Acosta,2
L.
Barbeyto,3 and
F.
Real2
Departamento de Biología Funcional y
Ciencias de la Salud, Facultad de Ciencias, Universidad de Vigo,
Campus de Ourense,1 and Laboratorio de
Microbiología, Hospital Cristal
Piñor,3 Ourense, and
Departamento de Patología Animal, Facultad de
Veterinaria, Universidad de Las Palmas de Gran Canaria, Las Palmas
de Gran Canaria,2 Spain
Received 28 January 1999/Returned for modification 5 June
1999/Accepted 21 August 1999
 |
ABSTRACT |
Hafnia alvei is a gram-negative facultatively anaerobic
bacillus that belongs to the family Enterobacteriaceae.
This organism is a causative agent of intestinal disorders and is found
in different environments. H. alvei has received increased
clinical attention as a cause of different infections in humans. This
study was performed to compare the MicroScan WalkAway automated
identification system in conjunction with the new MicroScan Combo
Negative type 1S panels with conventional biochemical methods for
identification of 21 H. alvei strains. The MicroScan
WalkAway system was found capable of correctly identifying 20 of the 21 strains tested.
 |
TEXT |
Hafnia alvei is a
gram-negative facultatively anaerobic bacillus that belongs in the
family Enterobacteriaceae. It is suspected to cause a
variety of intestinal disorders, including gastroenteritis (2, 21,
24, 28, 35). H. alvei has also been isolated from
various mammals (29), fish (10, 26), birds
(11, 20), soil, water (4, 28, 30), and a number
of foods (6, 8, 14, 23, 32). H. alvei possesses
several different virulence mechanisms, which are similar or identical
to those of other gram-negative enteropathogens (3). In
humans, H. alvei is a recognized cause of a number of
illnesses, including pneumonia (13), meningitis (17), abscesses (1), and septicemia
(16).
Recently, automated systems have been developed to identify
gram-negative bacteria (9, 15, 18, 19, 22, 27, 33), but the
reports about the evaluation of these systems did not include a large
number of H. alvei strains. The MicroScan WalkAway (Dade
MicroScan, Inc., Sacramento, Calif.) is an automated, commercially available system for rapid identification and susceptibility testing of
gram-negative bacilli and has received favorable reports relative to
the identification of these bacteria (5, 11, 15, 22, 25).
MicroScan has recently marketed MicroScan Combo Negative type 1S
panels. The panels are designed to identify to the species level
aerobic or anaerobic facultative gram-negative bacilli. The system uses
fluorogenic substrates and a pH indicator to detect bacterial enzymatic
activity. The purpose of this study was to evaluate the ability of the
MicroScan WalkAway system in conjunction with the new Combo Negative
type 1S panels to identify H. alvei strains.
Bacterial strains.
Twenty-one Hafnia alvei strains
were selected for testing (Table 1). The
strains were identified with the MicroScan WalkAway system and were
tested in parallel by standard reference procedures (31).
The strains were routinely cultured on Trypticase soy agar (TSA;
Cultimed) at 37°C for 24 h and stored on TSA slants at 4°C
under mineral oil and frozen at
70°C with 15% glycerol.
MicroScan panels.
Conventional MicroScan Negative
Combo type 1S panels were inoculated with the strains by the turbidity
standard technique. The panels were incubated for 24 h at 35°C
within the MicroScan WalkAway system. All procedures were performed
according to the manufacturer's directions (7).
Comparison of biochemical tests.
The following biochemical
tests were performed: D-glucose, sucrose,
D-sorbitol, raffinose, L-rhamnose,
L-arabinose, myoinositol, D-adonitol, and
melibiose; urease; hydrogen sulfide (H2S)
production; indole production; decarboxylation of lysine and ornithine;
arginine dihydrolase; tryptophan deaminase (TDA); esculin hydrolysis;
Voges-Proskauer (VP); utilization of citrate;
o-nitrophenyl-
-D-galactopyranoside (ONPG);
and OF-glucose.
Results from comparison of different assay systems in testing important
biochemical characteristics for the identification of H. alvei strains are listed in Table 2.
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|
TABLE 2.
Comparison between MicroScan WalkAway system and
conventional laboratory tests to evaluate important biochemical
characteristics for identification of H. alvei strains
|
|
The MicroScan identification patterns for the 20 strains correctly
identified as
H. alvei were positive for the fermentation
of
D-glucose and
L-arabinose, decarboxylation of
lysine and ornithine,
and utilization of citrate and OF-glucose and
were negative for
fermentation of sucrose,
D-sorbitol,
raffinose, myoinositol,
D-adonitol,
and melibiose; hydrogen
sulfide (H
2S) production; indole production;
and TDA; and
were variable for fermentation of
L-rhamnose, urease,
arginine dihydrolase, esculin, VP, and
ONPG.
The MicroScan WalkAway system was able to identify 20 of 21 of the
H. alvei strains tested (95%), and only 1 strain,
H. alvei 14-67, was misidentified as a rare biotype.
Strain 14-67 was negative
in the
L-rhamnose and urease
tests, the same as strain 11-69,
but in the case of strain 14-67, the
MicroScan system interpreted
the urease test as positive, which
produced a misidentification
as a rare biotype. In the case of strain
11-69, the urease test
was negative, and the final identification was
H. alvei, with
a probability of 99.9%.
Initially, the MicroScan system classified as a rare biotype strains
1967-82, F-4319, 30-65, and 7-68. The arginine test was
initially
interpreted as positive, but after a subsequent manual
reading of the
panels by specialized personnel in the center,
it was recorded as
negative. Therefore, the final identification
was
H. alvei,
with a probability of 99.9%.
Although
H. alvei infections are relatively rare, their
clinical importance is well documented. More accurate and reliable
methods with which to rapidly identify these organisms need to
be
developed. Some studies on the reliability of automated systems
have
shown good results, but insofar as the
H. alvei strains are
concerned, only small numbers were evaluated (
34,
36,
37),
with the unique exception of Kelly et al. (
12), who, using
38
Hafnia alvei strains, obtained results similar (92%) to
those
in our
study.
On the whole, the MicroScan WalkAway system in conjunction with the new
Combo Negative type 1S panels proved to be very useful
and reliable in
identifying
H. alvei strains of different origins.
This
system correctly identified 20 of the 21 strains tested in
this study
(95%). In 16 of 21 tests analyzed, the correlation
between the
MicroScan system and conventional tests was 100%.
Occasionally, the
erroneous test results were important enough
to result in a
misidentification. Discrepancies might be expected
because conventional
tests may represent standard tubed media
read after overnight
incubation, thus accepting some loss of precision
and accuracy for the
sake of convenience. Such procedures could
not represent appropriate
reference methods for evaluating automatic
systems. The low percentages
of correlation in some tests (urease,
23.8%; VP, 14.2%; citrate,
4.7%; and ONPG, 47.6%) did not seem
to affect the final
identification, although they should be considered
if other
gram-negative bacteria are
tested.
The arginine dihydrolase test presented interpretation difficulties.
This test caused a misidentification of one strain as
a rare biotype on
the MicroScan system. Therefore, it is recommended
that this test be
read manually in the event of a positive result
in the automatic
reading.
Generally, the
L-rhamnose test is positive for
H. alvei, although there are a few samples that are
L-rhamnose negative (strains
11-69 and 14-67). When the
L-rhamnose test is negative, special
attention should be
paid to the urease test, because a positive
urease result gives a rare
biotype in the MicroScan system, while
a positive
L-rhamnose result always gives the identification of
H. alvei, independent of the urease
test.
In conclusion, the results of this study confirm that the MicroScan
WalkAway system, in conjunction with the new MicroScan
Combo Negative
type 1S panels, is reliable for identification
of
H. alvei strains.
 |
ACKNOWLEDGMENTS |
We thank J. Michael Janda for critical review of the manuscript.
The assistance of the staff of the Clinical Microbiology Laboratory at
the Hospital Cristal Piñor Ourense is greatly appreciated. We
thank Cultimed-Spain for providing culture media.
L. A. Rodríguez is the recipient of a University of Vigo grant.
 |
FOOTNOTES |
*
Corresponding author. Mailing address: Área de
Microbiología, Departamento de Biología Funcional y
Ciencias de la Salud, Universidad de Vigo, Campus de Ourense, Las
Lagunas, 32004 Ourense, Spain. Phone: 988 387066. Fax: 988 387001. E-mail: lalopez{at}uvigo.es.
 |
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Journal of Clinical Microbiology, December 1999, p. 4186-4188, Vol. 37, No. 12
0095-1137/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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