Identification of New Influenza B Virus Variants by Multiplex Reverse Transcription-PCR and the Heteroduplex Mobility Assay

A quick genetic approach for the screening of influenza virus variants was developed in this laboratory (S. Zou, J. Clin. Microbiol. 35:2623–2627, 1997). It uses multiplex reverse transcription and multiplex PCR to amplify and differentiate the variable region of the hemagglutinin genes of different types and subtypes of influenza viruses. Variants within the same type or subtype are then identified by the heteroduplex mobility shift assay of the amplicons. The method was used to screen influenza virus isolates received from provincial laboratories during the 1996–1997 season and was able to identify new influenza B virus variants. Sequencing of the amplicons derived from the hemagglutinin gene of the identified variants and comparison with the vaccine strain B/Harbin/7/94 showed substitution rates of 2.26 to 2.55% at the nucleotide level and 4.26 to 4.68% at the amino acid level. The result further demonstrated that the approach provides a quick, sensitive, and reliable screening for influenza virus variants. It also suggested the necessity of close monitoring of influenza B virus isolates in the 1997–1998 season and critical evaluation of the reference strain for the type B influenza virus.