Simultaneous subtyping and pathotyping of the 2010–2011 South Korean HPAI outbreak strain by using a diagnostic microarray

The highly pathogenic avian influenza (HPAI) subtype H5N1 virus has caused continuous outbreaks in the poultry industry with devastating economic losses and is a severe threat to public health. In the present study, we developed a low-density microarray for the rapid detection and identification of avian influenza virus hemagglutinin subtypes H5, H7, and H9 and neuraminidase subtypes N1, N2, and N3. We evaluated this diagnostic microarray using 6 HPAI H5N1 clade 2.3.2 viruses that caused the South Korean outbreaks during the 2010–2011 winter season. Cy3-labeled DNA targets were generated by reverse transcription polymerase chain reaction (RT-PCR) using Cy3-labeled universal primers. The Cy3-labeled RTPCR products were then hybridized to the microarray. All subtypes and pathotypes were correctly determined and were found to be identical to the nucleotide sequencing results. This diagnostic microarray has enormous potential for the rapid subtyping and pathotyping of avian influenza viruses.