N.Naranbold,
National Influenza Laboratory,
National Center of Communicable Diseases, Mongolia
The 10th International Congress of the Asian Society of
Clinical Pathology and Laboratory Medicine
September 10-11, 2009 in Ulaanbaatar Mongolia.
Introduction
Rapid detection and identification of causal
agents is essential for timely actions to control
of outbreaks and successful clinical
management of the disease cases.
Introduction
However the traditional methods of virology
are time-consuming and costly, making them
less successful for practicising epidemiologists
and clinicians, especially to control shortduration infections like influenza.
Background
In this report we share with you our results in
the last 2 years with real-time Reversetranscriptase (r-RT) PCR for this purpose.
Background
In our laboratory, we have been introduced
real time PCR since 2008. The study aim is to
compare r-RT PCR and traditional method to
satisfy epidemiological and clinical needs. We
are trying to find out sensitive, rapid and
cheaper methods for the reliable detection of
A (H3, H1) and B influenza viruses.
Methods and Materials
• Samples
– 565 clinical nasal samples from patients with ILI collected in
ISSSs from Baganuur District, UB and Selenghe aimag in
2008/2009 cold season.
• r-RT-PCR
–
–
–
–
–
Instrument
RNA purification
Primer
Probe? !
Master-mix
(ABI 7300)
(QUIAGEN mini RNA kit)
(Invitrogen®A (H1), A(H3), B, M?
(TaqMan)
(Invitrogen)
• Traditional method
– Influenza virus isolation
– Serological tests
(MDCK cell culture)
(HA and HIT)
Traditional influenza
detection method
Specimen
collection
Virus Isolation
on MDCK cell
culture
~6-7days
Detection: HA
Subtyping: HIT
MDCK cell culture ready
for virus inoculation
MDCK cell culture with
virus CPE
r-RT-PCR influenza detection
method
Specimen
collection
RNA isolation
4-5 hours
r-RT-PCR
r-RT-PCR Thermal cycler
protocol
r-RT-PCR Results
Detection percentage
Duration
Expenses
Conclusion
r-RT-PCR
•6 times more sensitive
•30 times quicker
•2.5 times cheaper
Traditional method
(virus isolation on
cell culture)
for influenza virus detection
Conclusion
On the basis of the obtained results
r-RT_PCR is recommended for
routine influenza virus surveillance.