Guangxi Veterinary Research Institute, 51 You Ai North Road, Nanning, Guangxi 530001, People´s Republic of China.

A multiplex reverse transcriptase-polymerase chain reaction (mRT-PCR) was developed and optimized for the detection of type A influenza virus; the assay simultaneously differentiates avian H5, H7 and H9 hemagglutinin subtypes. Four sets of specific oligonucleotide primers were used in this test for type A influenza virus, H5, H7 and H9 heamagglutinin subtypes. The mRT-PCR DNA products were visualized by gel electrophoresis and consisted of fragments of 860bp for H5, 634bp for H7, 488bp for H9 hemagglutinin subtypes, and 244bp for type A influenza virus. The common set primers for type A influenza virus were able to amplify a 244bp DNA band for any of the other subtypes of AIV. The mRT-PCR assay developed in this study was found to be sensitive and specific. Detection limit for PCR-amplified DNA products was 100pg for the subtypes H5, H7, and H9 and 10pg for type A influenza virus in all subtypes. No specific amplification bands of the same sizes (860, 634 and 488bp) could be amplified for RNA of other influenza hemagglutinin subtypes, nor specific amplification bands of type A influenza (244bp) for other viral or bacterial pathogens.