Using clinical materials from experimentally infected poultry, we established an effective method for the preparation of viral RNA directly from tissue samples and eggs. Furthermore, our type A-specific matrix reverse transcription-polymerase chain reaction (RTPCR) test was improved, and an H7 subtype-specific nested RT-PCR, which includes the hemagglutinin cleavage site, was designed. Both RT-PCR systems proved to be as sensitive as virus isolation. In addition, the labeled H7 HA-nested PCR primers were suitable for sequencing of the PCR products. The RT-PCR amplification of viral RNA and sequencing of the PCR product allows for the sensitive and rapid differentiation between low-pathogenic and highly pathogenic avian influenza viruses.
