[Multiplex RT-PCR for the rapid detection of influenza virus types and subtypes].
OBJECTIVE
To establish a sensitive and specific multiplex RT-PCR(MRT-PCR) for the simultaneous detection of influenza virus types and subtypes.


METHODS
Primers were designed from highly conserved region of the hemagglutinin(HA) gene of influenza H1N1H3N2 and B virus and MRT-PCR was performed. Additional two pairs of primers were designed to determine the N1 and N2 subtypes of neuramidinase NA of influenza H1N1 and H3N2 virus.


RESULTS
The fragments of HA gene of all types of influenza virus were amplified and there was no cross reaction. The sensitivity of detection of influenza H1N1 and H3N2 virus was 0.10 TCID50/50 microliter by the second PCR and that was 0.01 TCID50/50 microliter for influenza B virus. The NA gene of influenza H1N1 and H3N2 virus was also amplified by this method.


CONCLUSIONS
The sensitivity of detection of influenza virus from clinical patients' throat washing specimens by MRT-PCR was higher than that by MDCK cell culture or egg embryo isolation. This method was highly sensitive and timely for detection of influenza virus types and subtypes.
Lu
Yi-yu
YY
Zhejiang Center for Disease Control and Prevention, Hangzhou 310009, China.
Yan
Ju-ying
JY
Mao
Hai-yan
HY
Sun
Yan
Y
Zhou
Min
M
Shi
Wen
W
D000818:Animals; D002460:Cell Line; D002642:Chick Embryo; D002645:Chickens; D017931:DNA Primers; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000302:isolation & purification; D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics / Q000302:isolation & purification; D009980:Influenza A virus / Q000235:genetics* / Q000302:isolation & purification; D005585:Influenza in Birds / Q000821:virology; D007251:Influenza, Human / Q000821:virology; D012367:RNA, Viral / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
D017931:DNA Primers; D012367:RNA, Viral