Inexpensive Test Detects H5N1 Infections Quickly
and Accurately
Scientists from the University of Colorado at Boulder and the
Centers for Disease Control and Prevention (CDC) have developed
an inexpensive “gene chip” test based on a single influenza virus
gene that could allow scientists to quickly identify flu viruses,
including avian influenza H5N1. The researchers used the MChip
to detect H5N1 in samples collected over a three-year period from
people and animals in geographically diverse locales. In tests
on 24 H5N1 viral isolates, the chip provided complete information
about virus type and subtype in 21 cases and gave no false positive
results, report the scientists. They say the MChip could provide
a significant advantage over available tests because it is based
on a single gene segment that mutates less often than the flu genes
typically used in diagnostic tests. As a result, the MChip may
not need to be updated as frequently to keep up with the changing
virus.
The research was led by University of Colorado scientist Kathy
L. Rowlen, Ph.D., and funded by the National Institute of Allergy
and Infectious Diseases (NIAID), part of the National Institutes
of Health. A paper describing the work, now available online, is
scheduled to appear in the December 15 issue of the American Chemical
Society’s journal Analytical Chemistry.
“Concerns about a possible influenza pandemic make it imperative
that we continue to devise reliable and easy-to-use diagnostic
tests for H5N1 that can be employed on-site where outbreaks are
suspected,” says NIAID Director Anthony S. Fauci, M.D. “The MChip
developed by Dr. Rowlen and her colleagues performed extremely
well in initial tests and has the potential to be a valuable tool
in global influenza surveillance efforts.”
The MChip has several advantages over the FluChip, a flu diagnostic
previously developed by the same research team, says Dr. Rowlen.
While the FluChip is based on three influenza genes — hemagglutinin
(HA), neuraminidase (NA) and matrix (M) — the MChip is based
on one gene segment. Unlike HA and NA, which mutate constantly
and thus are technically difficult to use to develop gene chip
diagnostic tests, the M gene segment mutates much less rapidly,
Dr. Rowlen explains. “The M gene segment is much less of a moving
target than the HA or NA gene. We believe that a test based on
this relatively unchanging gene segment will be more robust because
it will continue to provide accurate results even as the HA and
NA genes mutate over time. The work summarized in our paper strongly
supports that idea,” she says.
Another potential advantage is that the MChip would, for the first
time, create a way to simultaneously screen large numbers of flu
samples to learn both the type and subtype of virus present. Current
real-time tests provide information about the type of virus (type
A or B) in a sample, but additional tests must be run to determine
the virus subtype (for example, H5N1 subtype.)
Working in biosafety-level-3-enhanced labs in Atlanta, CDC scientists,
including Catherine B. Smith, M.S., extracted H5N1 genetic material
from virus samples derived from human, feline and multiple avian
hosts, including geese, chickens and ducks. The samples represented
infections that had occurred between 2003 and 2006 over a vast
geographic area, including Vietnam, Nigeria, Indonesia and Kazakhstan.
Six of the human viral isolates were taken from an Indonesian family
in which human-to-human H5N1 virus transmission was suspected.
The virus diversity in the samples is important, explains Dr. Rowlen,
because any diagnostic tool designed for eventual use on a rapidly
changing virus, such as H5N1, must be able to detect as many variants
as possible.
Dr. Rowlen and her colleagues tested the ability of the MChip
to correctly identify 24 different H5N1 viral isolates, and distinguish
those from seven non-H5N1 isolates. The MChip accurately identified
and gave complete subtype information (identifying the samples
as H5N1) for the 21 out of 24 strains of H5N1. Importantly, notes
Dr. Rowlen, the test gave no false positives, meaning that the
chip never indicated the presence of H5N1 when none was present.
Following exposure to a viral isolate, the MChip displays results
as a pattern of fluorescent spots. To automate the process of interpreting
this pattern — thus eliminating the possibility of human
error — the researchers developed an artificial neural network
trained to recognize the distinctive pattern indicative of H5N1.
Automating the interpretation of MChip results could allow it to
be used more readily by health workers at the site of possible
flu outbreaks, notes Dr. Rowlen.
“This new technology, once manufactured and distributed, could
have the potential to revolutionize the way laboratories test for
influenza,” says Nancy J. Cox, Ph.D., director of the CDC’s influenza
division. “The MChip could enable more scientists and physicians,
possibly even those working in remote places, to more quickly test
for H5N1 and to accurately identify the specific strain and its
features. This would greatly increase our ability to learn more
about the viruses causing illness and take the best steps to respond.”
The raw materials for the MChip cost less than 10 dollars, Dr.
Rowlen says, and discussions are under way to commercialize its
manufacture.
For more information on influenza see http://www3.niaid.nih.gov/news/focuson/flu.
Also visit http://www.PandemicFlu.gov for
one-stop access to U.S. Government information on avian and pandemic
flu.
NIAID is a component of the National Institutes of Health.
NIAID supports basic and applied research to prevent, diagnose
and treat infectious diseases such as HIV/AIDS and other sexually
transmitted infections, influenza, tuberculosis, malaria and
illness from potential agents of bioterrorism. NIAID also supports
research on basic immunology, transplantation and immune-related
disorders, including autoimmune diseases, asthma and allergies.
News releases, fact sheets and other NIAID materials are available
on the NIAID Web site at http://www.niaid.nih.gov.
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