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Comparative Evaluation of Four Enzyme Immunoassay
Tests versus Cell Culture for the Rapid
Diagnosis of Respiratory Syncytial Virus
Infection in a Pediatric Hospital
102nd A.S.M.May 19-23Salt Lake City UTAH
C-62
A.M. Freydiere 1, T. Goyard 1, C. Ploton 1, D.
Thouvenot 2, A. Rousson 1and F. Vandenesch 1 1
Hôpital Debrousse, Hospices Civils de Lyon, 2
Laboratoire deVirologie, Hospices Civils de Lyon,
Lyon, France.
E-mail am.freydiere_at_chu-lyon.fr
Abstract Rapid diagnosis of Respiratory
Syncytial Virus (RSV) impacts positively on
patient care by reducing hospital stays, and
decreasing the need for additional diagnostic
procedures. This study assesses the performance
of three rapid membrane enzyme immunoassay tests
(MEIA) and one ELISA test for RSV detection
compared with cell-culture considered as the
reference method. One hundred eighty six
endotracheal and nasopharyngeal aspirates from
hospitalized infants were tested prospectively,
53 with the DAKORapide VRS kit (J2L ELITECH,
France), 102 with the Directigen RSV kit (BECTON
DICKINSON, France), 103 with the RSV TESTPACK kit
(ABBOTT, France), and all the specimens were
tested both with ELISA Vidas test (bioMérieux,
France) and cell-culture. By using culture as the
gold standard, the sensitivities,
specificities, positive predictive values (PPV)
and negative predictive values (NPV) were 75,
67.8, 66.6, and 76 for DAKORapide VRS 80.6,
61.3, 81.8, and 59.4 for Directigen RSV 86.2,
77.7, 83.3, and 81.4 for RSV TESTPACK 87.2,
82.1, 85.5, and 84.1 for ELISA test. DAKORapide
VRS and Directigen RSV yielded 0.2 and 3.9 of
non-interpretable results, respectively, due to
difficulties in readings while the other tests
did not yield any non-interpretable
results. Retrospectively, 84 frozen endotracheal
and nasopharyngeal aspirates were simultaneously
tested with the MEIA tests. Compared to
cell-culture, the sensitivities, specificities,
PPV and NPV were 82.3, 60.6, 76.4, and 68.9 for
DAKORapide VRS 78.4, 66.6, 78.4, and 65.6 for
Directigen RSV 86.3, 63.6, 78.6, and 75 for
RSV TESTPACK. With frozen specimens DAKORapide
VRS showed a highest sensitivity than with fresh
specimens. Directigen RSV presented similar
results with either frozen or fresh specimens.
RSV TESTPACK achieved the best results among the
three MEIA tests and similar results to the ELISA
test. This test is rapid (20 min vs 2h 30 for
ELISA) and easy-to-perform it appears to be the
most convenient test for rapid diagnosis of RSV
in a microbiological laboratory.
Material and Methods Specimens. From October
1999 to April 2000 and from October 2000 to April
2001, 186 endotracheal aspirates or
nasopharyngeal aspirates were collected by
physicians from infants and children (from 1 to
18 months) with apparent viral respiratory tract
infections, admitted at Debrousse Hospital. 84 of
the 186 specimens were then frozen at -20C for
further testing by the three MEIA in
parallel. Reagents. The three MEIA evaluated
were the DAKORapide VRS kit (J2L ELITECH,
France), the Directigen RSV kit (BECTON
DICKINSON, France) and the RSV TESTPACK kit
(ABBOTT, France). Each MEIA was performed
following manufacturers instruction. The VIDAS
RSV ELISA test (bioMérieux) is an automated
qualita-tive test using the Enzyme Linked
Fluorescent Assay Technique. Cell Culture.
Samples are inoculated onto the following cell
lines LLC-MK2, MDCK, Vero, HEp-2 and MRC-5.
Cultures were performed on shell-vial with a
centrifugation after inoculation and incubation
at 34C with a 5 CO2 atmosphere. Cytopathic
effect was regularly checked for 10 days in HEp-2
and RSV and identified by immunofluorescence with
the RSV Direct IF reagent (bioMérieux) I)
Comparison of methods for the detection of the
RSV with cell culture, using fresh specimens. All
the clinical specimens received at the laboratory
were tested, prospectively, in real clinical
laboratory settings, at least by one of the three
MEIA, and were sent in parallel to the clinical
virology laboratory for testing by ELISA and cell
culture. Among the total of 186 specimens tested,
53 were tested by the DAKORapide VRS, 102 by the
Directigen RSV, 103 by the RSV TESTPACK and 186
by the VIDAS RSV ELISA. All the results were
compared to the cell culture. II) Comparative
evaluation of the three MEIA using frozen
clinical specimens. 84 frozen specimens were
retrospectively tested in parallel by the three
MEIA, in the same conditions. The results were
compared with the cell culture performed before
freezing of the specimens.
Results
I) Comparison of the three MEIA for RSV
detection with cell culture, using fresh specimens
Results are presented in Table 1 a and 1 b.
These findings, in accordance with other studies
(7, 8, 12) are most likely due to insensitive RSV
culture methods and/or problems with transport of
specimens to the laboratory since RSV is a
relatively labile virus and isolation is
optimized if specimens are inoculated rapidly.
On the contrary, in some other cases, only cell
culture was positive.
Table 1 a. Comparison of the three MEIA for the
detection of the RSV with cell culture, using
fresh specimens
Number of specimens
Cell culture method
Positive
Negative
Positive
Negative
Positive
Negative
Positive
Negative
II) Comparative evaluation of the three MEIA,
using frozen clinical specimens.
Non-interpretable result
84 frozen specimens were tested in parallel with
the three MEIA in the same conditions.
DAKORapide VRS and Directigen RSV yielded 0.2 and
3.9 of non-interpretable results,
respecti-vely, due to difficulties in readings.
Table 2. Sensitivity, Specificity, PPV and NPV
for the three MEIA using 84 frozen specimens and
the cell culture as reference method.
Some authors used freeze-thawing to increase the
sensitivity of EIAs (4, 10). In this study only
the DAKORapide VRS showed a higher sensitivity
after freezing (82.3 versus 75 ). Again, RSV
TESTPACK had the highest sensitivity and overall
similar results in specificity in comparison with
the two other MEIA. The commercially-available
MEIA tests for RSV detection are notable because
of their convenience and ease-to perform.
Moreover, the use of these rapid methods have
been demonstrated to have clinical and financial
benefits (1, 11). Recent studies have
demonstrated that genetical methods are more
sensitive than both cell culture and
immunological methods (2, 5, 6), however, until
real-time genetical methods becomes a reality,
direct specimen testing such as immunoenzymatic
methods is likely to remain the most rapid,
sensitive and specific methodology for detection
of RVS in clinical specimens.
Introduction Respiratory syncytial virus (RSV)
represents the single most important cause of
serious lower respiratory tract infection being
involved, especially bronchiolitis,
tracheobronchitis, and pneumoniae, among infants
and children in all parts of the world (3). It is
the primary cause of hospitalization in the first
year of life.The rapid identification of
respiratory pathogens is of great importance for
the prevention of nosocomial spread, for
monitoring infected patients and for improving
clinical management. Since the virology
laboratory in not located within our pediatric
hospital, we determined whether rapid membrane
enzyme immunoassay tests (MEIA) might be used in
our local bacteriological laboratory and which of
the different commercialized tests yielded the
best performance. This study assesses the
performance of three rapid MEIA and one ELISA
test (VIDAS RSV test) for RSV detection compared
with cell-culture considered as the reference
method by using nasopharyngeal aspirates obtained
from pediatric infants with suspected viral lower
respiratory tract infections.
Amongst the three MEIA, the RSV TESTPACK had the
highest sensitivity and specificity. In previous
studies, the sensitivity of RSV TESTPACK compared
with culture varies from 83 to 96 and
specificity from 84 to 97 according to the
authors (3, 9). The specificity of the three
MEIA increased when combining, for the reference
method, the results obtained by cell culture and
VIDAS RSV ELISA test.
Conclusion
The results of this comparative study, allowed us
to use the rapid RSV TESTPACK from ABBOTT to
screen incoming clinical specimens to identify
any positive results in 20 minutes. All the
negative specimens are sent to the clinical
virology laboratory for further testing using
more sensitive diagnostic tests.
Acknowledgements We thank the technicians of the
two laboratories for their technical assistance,
ABBOTT, BECTON DICKINSON, and J2L ELITECH for
providing us their respective reagents, and Anne
Frangin for her valuable help in producing this
poster.