Identification and subtyping of avian influenza viruses by reverse transcription-PCR.
Avian influenza viruses have 15 different hemagglutinin (HA) subtypes (H1-H15). We report a procedure for the identification and HA-subtyping of avian influenza virus by reverse transcription-PCR (RT-PCR). The avian influenza virus is identified by RT-PCR using a set of primers specific to the nucleoprotein (NP) gene of avian influenza virus. The HA-subtypes of avian influenza virus were determined by running simultaneously 15 RT-PCR reactions, each using a set of primers specific to one HA-subtype. For a single virus strain or isolate, only one of the 15 RT-PCR reactions will give a product of expected size, and thus the HA-subtype of the virus is determined. The result of HA-subtyping was then confirmed by sequence analysis of the PCR product. A total of 80 strains or isolates of avian influenza viruses were subtyped by this RT-PCR procedure, and the result of RT-PCR gave an excellent (100%) correlation with the result of the conventional serological method. The RT-PCR procedure we developed is rapid and sensitive, and could be used for the identification and HA-subtyping of avian influenza virus in organ homogenates.
Lee
M S
MS
Department of Veterinary Medicine, National Chung Hsing University, Taichung, Taiwan.
Chang
P C
PC
Shien
J H
JH
Cheng
M C
MC
Shieh
H K
HK
D000818:Animals; D001717:Birds / Q000821:virology*; D017931:DNA Primers / Q000235:genetics; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics; D006801:Humans; D009980:Influenza A virus / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D007251:Influenza, Human / Q000821:virology; D009698:Nucleoproteins / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D017931:DNA Primers; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D009698:Nucleoproteins; D012367:RNA, Viral