Pathology Associates of the Roaring Fork Valley
Volume 17, Number 2
Frank Holmes, M.D.
Laboratory Diagnosis of Novel Influenza A (H1N1)
Robert Macaulay, M.D.
and the Limitations of Rapid Influenza Diagnostic Tests
Jerry Steinbrecher, M.D.
The sudden appearance and worldwide tory testing. These enzyme immunoassays
spread in early 2009 of pandemic novel (EIAs) are inexpensive, easy to use and offer
influenza A (H1N1), also known as swine-ori- rapid turnaround time, often less than thirty
Aspen Valley Hospital
gin influenza A (S-OIA), has necessitated the minutes. And, while very specific for influenza
Grand River Hospital
Pioneers Medical Center optimization of laboratory diagnosis of when positive, RIDTs have limited sensitivity.
influenza and, importantly, highlighted the A recent CDC study of RIDTs revealed that,
important limitations of widely used commer- while these assays are certainly capable of
Vail Valley Medical Center
cial rapid influenza diagnostic tests (RIDTs). detecting novel influenza A (H1N1) from res-
Valley View Hospital
Within days of identifying the novel strain, piratory specimens, overall sensitivity is low,
the CDC had sequenced the virus’s genome ranging from 40 to 69%, declining substantial-
and developed a real-time polymerase chain ly as viral levels decrease1. Another study
reaction (RT-PCR) assay to detect the virus. revealed sensitivity as low as 10%2.
Presently, this RT-PCR assay, which has An additional important limitation is the
received emergency use authorization from inability of rapid EIAs to distinguish between
the FDA, is the only means of differentiating seasonal influenza and novel influenza H1N1
novel H1N1 from seasonal influenza strains which, as noted above, is dependent on RT-
and thus confirming a diagnosis of “swine flu.” PCR assays.
RT-PCR is the most sensitive assay present- Based in part on these performance charac-
ly available for the detection of influenza virus teristics, the CDC in July 2009 issued interim
in clinical specimens. Viral culture is also high- guidelines for the detection of novel H1N1
ly sensitive but, requiring up to seven days for virus (see Table). Note that, given the excellent
results, is not widely employed. As the vast specificity of most RIDTs, a positive result can
“A negative rapid
majority of clinical microbiology laboratories be used in making treatment decisions while,
are not presently capable of offering PCR or due to limited sensitivity, a negative result
does not rule viral culture, RIDTs are the mainstay of labora- Continued on page 2
Diagnostic Algorithm for H1N1 Virus3
RIDT (+) for Flu A RIDT (-) for Flu A
Interpretation Influenza A virus infection likely. Cannot rule out influenza
Could be novel H1N1, seasonal virus infection.
H1N1, H3N2, or, rarely, an
influenza A virus of animal origin.
Suggested Response Treat with antiviral agents, if appropri- Use clinical symptoms, severity,
ate. Consider additional testing and underlying disease to decide if
to determine Influenza A subtype. antiviral treatment is appropriate.
Consider if empiric antibiotic Do not use a negative test to send
therapy for co-infection is indicated. a symptomatic child back to
school, to rule out an institutional
outbreak, or to dictate infection
control measures. Consider
whether further influenza specific
testing (i.e., RT-PCR) is necessary.
Consider whether empiric antibiot-
ic therapy for co-infections is
does not rule out infection. The sensitivity of clinical specimen obtained. While a variety of
these rapid assays is dependent both on the respiratory specimens, including nasopharyn-
nature of the sample and the patient from whom geal and throat swabs, have been used with
it is collected. Influenza virus is first detected in RIDTs, nasopharyngeal aspirate is considered to
“Nasopharyngeal respiratory secretions immediately before the be the sample of choice4.
onset of illness; viral shedding rapidly increases In the not too distant future, we expect to
aspirate is and remains elevated for 24 to 48 hours, then have available, locally, PCR-based assays for
rapidly declines to low levels and is usually influenza. These assays, currently undergoing
undetectable after 5 to 10 days. Rapid diagnosis FDA evaluation, promise to render moot the
considered to be
of influenza is thus best accomplished during inherent problems with current rapid enzyme
the first 2 to 3 days of illness when viral shed- immunoassays.
the specimen of
ding is maximal. -RMM
The sensitivity of rapid tests tends to be
choice for rapid
lower in adults and older patients due to 1 MMWR August 7, 2009 58(30); 826-829.
acquired immunologic factors; conversely, due 2 Ginocchio, CG et al. Journal of Clinical Virology
to the relative lack of immunity and correspon- 45(2009):191-195.
ding prolonged shedding of high levels of virus, 3 Clinical Laboratory News September 2009.
4 Henry’s Diagnosis and Clinical Management by
RIDTs are more sensitive in children. Test sensi-
Laboratory Methods (21st Ed.): pp. 981-2.
tivity is also dependent on the quality of the